Novel compounds having antiviral activity

ABSTRACT

In order to provide the anti-retrovirus active compound with low anti-coagulant action and low cytotoxicity, compounds comprising glycoside or the salt thereof wherein lipid is linked to position 2 of sialic acid having all hydroxyl groups at positions 4, 7, 8 and 9 completely sulfated, or KDN (2-keto-3-deoxy-D-glycero-2-nononic acid) having all hydroxyl groups at positions 4, 5, 7, 8 and 9 completely sulfated are provided.

BACKGROUND OF THE INVENTION

[0001] 1. Field of the Invention

[0002] This invention relates to a novel compound with the anti-viralactivity and a drug and anti-viral agent containing said compound as theeffective ingredient, more specifically relates to a medicine andanti-HIV active compound comprising the sulfated nonulonic acid.

[0003] 2. Description of the Related Arts

[0004] Azidothymidine (AZT) and dideoxy inosine (DDI) have been used asthe AIDS remedy. These agents are to inhibit the reverse transcriptaseof HIV, bringing about apothanasic effect to patients, while posingproblems that the chronic administration of AZT causes myelopathy andthat the DDI administration results in side reactions such as acutepancreatitis and peripheral neuropathy. Furthermore, the use of eitherdrug eventually results in the generation of virus resistant to thesedrugs.

[0005] Recently, the sulfated polysaccharide has been expected as apromising AIDS remedy. It is well known that polysulfated compounds suchas dextransulfate (I) (Japanese Patent Laid-Open Publication No.Sho63-45233), polyvinyl alcohol sulfate (II) (Antimicrob. AgentsChemother. 34, 134-138 (1990)), oligosaccharide sulfate (Japanese PatentLaid-Open Publication No. Hei2-304025) inhibit the proliferation of HIV.These compounds are produced simply by binding the sulfate group topolysaccharides, oligosaccharides or organic polymeric molecules.

[0006] In addition, the sulfated modified cyclodextrin (III) iscyclodextrin to which lipid-soluble groups such as aryl-, alkyl-groupsare introduced, and also has the proliferation inhibiting activityagainst retroviruses, HIV in particular (Japanese Patent Laid-OpenPublication No. Hei4-136001).

[0007] In addition, as a virucide comprising the sulfated polysaccharideas the active ingredient is disclosed the one using an acyl derivativeof a sulfated oligosaccharide glycoside (Japanese Patent Laid-OpenPublication No. Hei6-256373).

[0008] At first, it was conceived that the virucidic activity ofsulfated polysaccharides was due to their reverse transcriptaseinhibiting activities, and now it is thought to be due to theinteraction between the sulfated polysaccharide and the coat proteingp120 of HIV.

[0009] However, since sulfated polysaccharides generally have theinhibitory activity for the blood coagulation system (anticoagulantactivity), they have not been accepted as the suitable medicine.

DISCLOSURE OF THE INVENTION

[0010] This invention has been developed in view of the above-mentionedproblems, aiming at providing the anti-viral compound with a relativelyweak anti-blood coagulating activity and a low cytotoxicity, especiallythe one with the anti-HIV activity.

[0011] The present inventors have actively pursued the study to resolvethe above-mentioned problems, establishing that the following compoundsare useful for attaining the above-described purpose and completing thepresent invention.

[0012] (1) A compound having all hydroxyl groups of the sugar moiety ofglycoside comprising monosaccharide-lipid sulfated or the salt thereof,wherein said lipid moiety is bound to the anomeric position of saidsugar moiety.

[0013] (2) The compound or the salt thereof according to (1), whereinsaid sugar and lipid moieties are connected by the O-glycosidic linkageor the S-glycosidic linkage.

[0014] (3) The compound or the salt thereof according to (2), whereinsaid lipid is linear, and said linear lipid has a branched structure.

[0015] (4) A compound or the salt thereof, wherein all hydroxyl groupsof nonuloic acid moiety of the glycoside comprising amonosaccharide-lipid with said lipid bound to the anomeric position ofnonuroic acid derivative are sulfated.

[0016] (5) A compound or the salt thereof, wherein all hydroxyl groupsof sialic acid moiety or KDN moiety of the glycoside comprising amonosaccaride-lipid with said lipid bound to the anomeric position ofsialic acid or KDN are sulfated.

[0017] (6) The compounds or the salts thereof according to (5), whereinthe binding of said sialic acid moiety or KDN moiety to the lipid moietyis either the O-glycosidic of S-glycosidic linkage at position 2 of saidmoiety or the amidic linkage at position 1 of said residue.

[0018] (7) The compound or the salt thereof according to (6), whereinsaid lipid is a linear lipid, and this lipid has a branched structure.

[0019] (8) The compound or the salt thereof according to (7), whereinsaid branched chain is localized at position 2 of the main chain of saidlipid moiety.

[0020] (9) The compound or the salt thereof according to (8), whereinsaid lipid moiety has a forked two chain structure due to saidbranching.

[0021] (10) The compound or the salt thereof according to (9), whereinsaid lipid moiety has an alkyl group with the skeleton-forming carbonatoms from 1 to 4 at said branching site.

[0022] (11) The compound or the salt thereof according to (9) or (10),wherein the total number of said lipid skeleton-forming atoms is from 22to 60.

[0023] (12) The compound or the salt thereof according to (11), whereinsaid branched chain comprises a carbon-carbon unsaturated bond.

[0024] (13) The compound or the salt thereof according to (11), whereinsaid branched chain is linear.

[0025] (14) The compound or the salt thereof according to (11), whereinsaid branched chain has an ester linkage or ether linkage, respectively.

[0026] (15) The compound or the salt thereof according to (14), whereinsaid ester linkage or ether linkage is localized at positions 1 or 2 ofsaid branched chain.

[0027] (16) The compound or the salt thereof according to (11), whereinthe number of skeleton-forming atoms is from 10 to 28 per one branchedchain.

[0028] (17) The compound or the salt thereof according to (16), whereinthe number of skeleton-forming atoms is from 18 to 26 per one branchedchain.

[0029] (18) The compound or the salt thereof according to (17), whereinthe number of skeleton-forming atoms is from 24 per one branched chain.

[0030] (19) The compound or the salt thereof according to (18), whereinsaid forked branched chains are of the same length, respectively.

[0031] (20) The compound or the salt thereof according to (19), whereinsaid forked branched chains are of the same, respectively.

[0032] (21) The compound or the salt thereof according to (20), whereinsaid branched chain has the ester bond or the ether linkage at itsposition 1 or 2.

[0033] (22) The compound or the salt thereof according to (21), whereinsaid branched chain is linear.

[0034] (23) A medicine comprising the compound according to anyone of(1) to (3) in a pharmaceutically effective dosage.

[0035] (24) An antiviral drug containing the compound according toanyone of (1) to (3) in a pharmaceutically effective dose.

[0036] (25) An anti-HIV drug containing the compound according to anyoneof (1) to (3) in a pharmaceutically effective dose.

[0037] (26) A drug containing the compound according to anyone of (1) to(21) in a pharmaceutically effective dose.

[0038] (27) An antiviral drug containing the compound according toanyone of (1) to (21) in a pharmaceutically effective dose.

[0039] (28) An anti-HIV drug containing the compound according to anyoneof (1) to (21) in a pharmaceutically effective dose.

[0040] Drugs (claims 23 and 26), furthermore, antiviral drugs (claims 24and 27), especially anti-HIV drugs (claims 24 and 27) containingcompounds according to any one of the above-described (1) to (22) arewithin the scope of the present invention. However, since drugs,anti-viral drugs and anti-HIV drugs containing the compounds accordingto any one of the above-described (5) to (22) in pharmaceuticallyeffective doses have lower anti-coagulant activity as well as lowerbiological toxicity than those containing compounds according to any oneof the above-described (1) to (3), the former group of drugs arepharmaceutically useful. It has been confirmed that, of theabove-described compounds, particularly, those having 24skeleton-forming atoms per one branched chain (24), furthermore, each ofsaid branched chains of which has the same structure, is linear and hasthe ether bond at its position 1 or 2, are pharmaceutically useful(examples 12, 13, 29, 50 and 59).

[0041] And, the usage of drugs, anti-viral drugs and anti-HIV drugscontaining the compound according to any one of the above-described (1)and (22) as the drugs, anti-viral drugs and anti-HIV drugs, and themethod for treating the particular diseases (viral disease and acquiredimmune deficiency syndrome caused by HIV) are also within the scope ofthe present invention.

BEST MODE FOR CARRYING OUT THE INVENTION

[0042] Definition of the Terms

[0043] As used in this specification, by the term “nonuloic acid” ismeant the same nonuloic acid as used generally, an acidic carbohydratehaving a carboxyl group at its position 1 and 9 carbon atoms.Accordingly, “nonuloic acid derivatives” used in this specificationinclude neuraminic acid(5-amino-3,5-dideoxy-D-glycero-D-galacto-nonuloic acid) and neuraminicacid derivatives. Since the later-described “sialic acid” is an acylderivative of neuraminic acid, it is also included in “nonuloic acidderivatives”.

[0044] By “KDN” used in this specification is meant2-keto-3-deoxy-D-glycero-2-nononic acid. And, by “sialic acid used inthis specification is meant the generic name for a series of derivativesof substances having the neuraminic acid as the basic structure (YasuoInoue, Seitai Bunsi no Kagaku, Tositu no Kagaku (Chemistry ofBiomolecules 1, Chemistry of Carbohydrates), p80-81, Baifukan), and acylderivatives, more specifically, such as N-acetylneuramic acid andN-glycolylneuramic acid are also included therein.

[0045] By “forked two chains” [for example, (9), claim 9] is meant astructure constructed with two chains having more than 7skeleton-forming atoms. Accordingly, alkyl groups according to claim 10(10), wherein the total number of their skeleton-forming atoms is 1 to4, are not included in “chain” constructing “forked two chains”.

[0046] By “skeleton-forming atoms” is meant atoms constructing theskeleton of the chain, including carbon atom, oxygen atom, nitrogenatom, sulfur atom, etc. However, monovalent atom, such as hydrogen atom,is not included in the “skeleton-forming atoms”, because it cannotconstruct the skeleton portion of the chain.

[0047] By “salt” is meant the sodium and potassium salts which arerequired to neutralize the intramolecular carboxylic acid and sulfonicacid, to which cations bind so as not to decrease the biologicalactivity. As to the cations bound to said acids, any cations which donot lower the biological activity of the compounds related to thepresent invention may be used.

[0048] Sugars

[0049] In principle, the compound related to this invention is aglycosidic compound and the salt thereof comprising a sugar-lipid withsaid lipid bound to the anomeric carbon of said sugar, wherein hydroxylgroups of said sugar moiety are all sulfated, having an excellentanti-retroviral activity [the above-described (1), claim 1]. Althoughthere have been hitherto found the sulfated sugars with theanti-retroviral activity, the glycoside comprising a sugar-lipid withsulfate group introduced and having the anti-retroviral activity has notbeen discovered. In this respect, this invention is valuable in findingthat a glycoside comprising a sugar-lipid with the sulfate groups fully(100%) introduced or the derivative thereof has an excellent anti-viralactivity.

[0050] When the sugar moiety of said glycoside is sialic acid or KDN,the glycoside not only has the strong anti-viral activity, but alsolower cytotoxicity, and can preferably achieve the main purpose asmedicine [the above-described (5) to (22), (26) to (28), and claims 5 to22 and claims 26 to 28]. And, in this case, the hydroxyl groups of thesugar moiety of the compound related to the present invention to be allsulfated to form the wholly sulfated nonulonic acid [the above-described(4), and claim 4] are at positions 4, 7, 8 and 9 when nonulonic acid isN-acetylneuramic acid, and the glycolyl hydroxyl group at position 5, inaddition to the hydroxyl groups at the positions 4, 7, 8 and 9. In thecase wherein nonulonic acid is KDN, all the hydroxyl groups at positions4, 5, 7, 8 and 9 are sulfated.

[0051] The bond between the monosaccharide moiety and lipid moiety ofthe compound related to the present invention can be of any type.Accordingly, the bond between the monosaccharide and lipid may be notonly the O-glycosidic linkage but also S-glycosidic linkage.Furthermore, in the case where the sugar moiety of the compound relatedto the present invention is nonuloic acid, the amide linkage and esterlinkage can be formed using the carboxyl group at position 1, inaddition to a glycosidic linkage with the carbon atom at position 2. Thebond between monosaccharide and lipid in the compound related to thepresent invention can be such amide linkage and ester linkage.Therefore, although, by “glycoside” is generally meant a compoundwherein the monosaccharide and lipid moieties are linked by a glycosidiclinkage, by “glycoside” in this specification is meant a compoundwherein the monosacchharide and lipid are bonded not only in aglycosidic linkage but also in amide and ester linkages. Accordingly,not only compounds having a glycosidic linkage at position 1 ofmonosaccharide such as glucose or position 2 of nonulonic acid but alsothose having an amide linkage or ester linkage at position 1 ofnonulonic acid are also in the scope of “glycoside” of the presentinvention.

[0052] However, in consideration of the overall manufacturing easinessand biological activity of these compounds, glycosides with theO-glycosidic, S-glycosidic, and amide linkages are preferred.

[0053] Lipids

[0054] By “lipid” in the compound related to this invention is meant thelipid in a broad sense including steroid, carotinoid, terpenoid, etc.,and conceptionally even the compound such as cholesterol. However, thecompound related to this invention is preferably a linear lipid, whichfurther preferably has the branched chain structure [the above-described(7), and claim 7]. The branch can be two-forked or three-forked, locatedat position 2 of the main chain of the lipid moiety. As a result, thelipid is preferably two-forked at the β position (β position withrespect to the sugar moiety) of said lipid moiety [the above-described(8) and (9), and claims 8 and 9]. Furthermore, this branching site mayhave alkyl group with the skeleton-forming atoms 1 to 4 [theabove-described (10), and claim 10]. “Alkyl group with theskeleton-forming atoms 1 to 4” herein cited includes, for example,methyl group, ethyl group, propyl group, etc.

[0055] The above-described two-forked chain can be a hydrocarbon chain,which can include heteroatoms such as oxygen, nitrogen, sulfur, etc.Furthermore, regardless of the species of component atoms, the totalnumber of the skeleton-forming atoms of lipid is preferably 22 to 60[the above-described (11), and claim 11]. In addition, theabove-described two-forked chains can have unsaturated bond betweencarbon atoms, respectively [the above-described (12), and claim 12].Also, although the above-described two-forked chain can be furtherbranched, they are preferably linear [the above-described (13), andclaim 13].

[0056] In the case where the above-described two-forked chain containsheteroatoms as the component atom, each branched chain preferablycontains an ester bond or ether bond [the above-described (14), andclaim 14], furthermore, said ester bond or ether bond is preferablylocalized at position 1 or 2 of said branched chain [the above-described(15), and claim 15].

[0057] Herein, when the ester bond or ether bond is present at position1 of the branched chain, the compound related to this invention willbecome the sulfated derivative of sialoglycerolipid with the excellentanti-reteroviral activity (In this connection, in the case of ether bondpresent, said compound will be an alkyl glycerol wherein a long-chainalcohol is linked to the glycerol residue, and in the case of ester bondpresent, said compound will be amyl glycerol.). Furthermore, in the casewhere ester bond or ether bond is located at position 2 of the branchedchain, the glycerol residue in the glycerol area is suitably modified tobecome pseud-glycerol.

[0058] As to the length of branched chains, the number ofskeleton-forming atom is preferably 10 to 28 [the above-described (18),and claim 16], more preferably 18 to 26 [the above-described (17), andclaim 17], most preferably 24 [the above-described (18), and claim 18].In addition, forked-chains can be of different lengths, but preferablyof the same length [the above-described (19, and claim 19], mostpreferably of the same structure comprised of the same component atoms[the above-described (20), and claim 20]

[0059] Method of Administration

[0060] When compounds related to the present invention are used astherapeutics, they are administered singly or in combination withpharmaceutically acceptable medical carriers, either organic orinorganic and either solid or liquid. Their compositions are determinedaccording to the solubility, chemical property, administration route andschedule of compounds.

[0061] Compounds related to this invention can be administered by anysuitable desired administration routes. More specifically, compoundsrelated to this invention can be administered intraperitoneally,subcutaneously, percutaneously, intravenously or intra-arterially, andlocally injected in the case of animals, and intravenously,intra-arterially, by local injection, intraperitoneally/intrapleurally,orally, subcutaneously, intramuscularly, sublingually, percutaneously,inhalationally or rectally in the case of humans.

[0062] Dosage Form

[0063] When compounds related to the present invention are administeredas the drug, they can be administered, according to the method andpurpose of their administration, in the form of injection, suspension,tablet, granule, powder, capsule, ointment, cream, suppository, tape,etc. For preparing these drugs, solvent, solubilizing agent, isotonizingagent, preservative, anti-oxidant, excipient, binder, lubricant,stabilizer, etc. can be added.

[0064] Solvents are exemplified, for example, by water, physiologicalsodium chloride solution, etc.; solubilizing agents, for example, byethanol, polysorbates, chromophore, etc.; excipients, for example, bylactic acid, sucrose, starch, cellulose, crystalline cellulose, dextrin,mannitol, maltose, kaolin, calcium hydrogenphosphate, light anhydroussilicic acid, calcium carbonate, etc.; binders, for example, by starch,polyvinylpyrrolidone, hydroxypropyl cellulose, ethyl cellulose,carboxymethyl cellulose, gum arabic, etc.; disintegrators, for example,by starch, calcium carboxymethyl cellulose, etc.; lubricants, forexample, by magnesium stearate, talc, hydrogenated oil, etc.;stabilizers, for example, by lactose, mannitol, maltose, polysorbates,macrogols, polyoxyethylene hydrogenated castor oil, etc. Furthermore,glycerol, dimethylacetamide, 70% sodium lactate, surfactant, basicsubstances (such as sodium hydroxide, ethylenediamine, ethanolamine,sodium carbonate, arginine, meglumine, tris-aminomethane) may be added,if needed. Using these components, compounds related to this inventioncan be prepared to the dosage forms such as injections, tablets,granules, capsules, etc.

[0065] Compounds related to this invention can be administered orally indosage forms such as granule, fine granule, powder, tablet, heavy syrup,soft capsule, syrup, emulsion, suspension, liposome, liquid preparation,etc. Liquid preparations such as emulsion, syrup, suspension, liquiddrug, etc. for oral administration comprise generally used inertdiluents, for example, water or vegetable oil. These preparations canalso include supplements, for example, moistening agents, suspendingagents, sweetening agents, aromatics, coloring matters, preservatives,etc., in addition to these inert diluents. Liquid preparations may beenveloped in capsules made of absorbable materials such as gelatin.

[0066] For administration by intravenous, intramuscular and subcutaneousinjections, compounds may be made in powder form for injection, andprepared for injection prior to use. Solvents or suspending agents forpreparing drugs for parenteral administration, that is, injections, etc.are exemplified, for example, by water, propylene glycol, polyethyleneglycol, benzylalcohol, ethyl oleate, lecithin, etc. Preparation ofpharmaceutics can be carried out according to the conventional method.

[0067] Treatment

[0068] It is desirable that clinical dosages are appropriately increasedand/or decreased according to the age, symptom, presence or absence ofsimultaneous administration of other drugs. The daily dosage ofcompounds related to this invention may be administered once daily, orin two or three divided portions at appropriate intervals, orintermittently. In view of results of animal experiments and variousconditions, dosages of compounds related to this invention aredetermined such that the total dosage does not exceed a certain limitregardless of whether they are administered once or repeatedly. Needlessto say that the optimum dosage is varied according to the method ofadministration, conditions of patients or animals to be treated such asage, body weight, sex, susceptibility, food (diet), time ofadministration, drugs in joint use, patient's conditions, severeness ofsymptoms, etc. In addition, the optimal dosage and administrationfrequency under the certain conditions must be determined by thesuitable usage determination test performed by a medical specialistaccording to the above-describe guideline.

BRIEF DESCRIPTION OF THE DRAWINGS

[0069]FIG. 1 is a graphic representation of the dependency of biologicalactivity of two-forked alkylglycerol with a glycosidic linkage on thechain length

[0070]FIG. 2 is a diagram showing the reaction pathway from Examples 1to 6.

[0071]FIG. 3 is a diagram showing the reaction pathway from Examples 7to 12.

[0072]FIG. 4 is a diagram showing the reaction pathway from Examples 13to 17.

[0073]FIG. 5 is a diagram showing the reaction pathway from Examples 18to 21.

[0074]FIG. 6 is a diagram showing the reaction pathway from Examples 22to 25.

[0075]FIG. 7 is a diagram showing the reaction pathway from Examples 26to 29.

[0076]FIG. 8 is a diagram showing the reaction pathway from Examples 30to 34.

[0077]FIG. 9 is a diagram showing the reaction pathway from Examples 35to 36.

[0078]FIG. 10 is a diagram showing the reaction pathway from Examples 37to 41.

[0079]FIG. 11 is a diagram showing the reaction pathway from Examples 42to 45.

[0080]FIG. 12 is a diagram showing the reaction pathway from Examples 46to 51.

[0081]FIG. 13 is a diagram showing the reaction pathway from Examples 52to 56.

[0082]FIG. 14 is a diagram showing the reaction pathway from Examples 57to 60B.

[0083]FIG. 15 is a diagram showing the reaction pathway from Examples60C to 63.

[0084]FIG. 16 is a diagram showing the reaction pathway from Examples 64to 68.

EXAMPLES

[0085] Synthesis of Compounds

Example 1 Sodium[methyl 5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosid]onate

[0086] Methyl5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosidonate[Chem. Ber., 99, 611 (1966)] (200 mg, 0.58 mmol) and sulfurtrioxide-trimethylamine complex (1600 mg, 11.5 mmol) were stirred inanhydrous dimethylformamide (5.8 ml) under the argon atmosphere at80-85° C. for 2 h. The reaction mixture was purified directly by silicagel column chromatography (gel 209 g, chloroform/methanol/water, 5:4:1),and further treated with Dowex 50W-X8 (Na form) resin. The product wasfurther purified by gel chromatography (Sephadex G-25, 400 ml, water) toobtain the title compound (359 mg, 82%) as white solid.

[0087]¹H-NMR (D₂O) δ: 3.37 (s, 3H, OCH₃), 2.88 (dd, 1H, J=4.0, 11.7 Hz,H-3eq), 1.97 (s, 3H, NAc), 1.77 (t, 1H, J=12.1 Hz, H-3ax).

Example 2 Sodium[methyl 5-acetamido-3,5-dideoxy4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-β-D-galacto-2-nonulopyranosid]onate

[0088] Methyl5-acetamido-3,5-dideoxy-D-glycero-β-D-galacto-2-nonulopyranosidonate[Chem. Ber., 99, 611 (1966)] (200 mg) was reacted by the generalprocedure according to Example 1 to obtain the title compound (218 mg,50%) as white solid.

[0089]¹H-NMR (D₂O) δ: 3.33 (s, 3H, OCH₃), 2.57 (dd, 1H, J=5.1, 13.2 Hz,H-3eq), 1.96 (s, 3H, NAc), 1.90 (t, 1H, J=13.2 Hz, H-3ax).

Example 3 3-O-[Sodium{5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosyl}onate]-1,2-di-O-hexyl-Sn-glycerol

[0090]3-O-[Sodium(5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-hexyl-Sn-glycerol[Japanese Patent Laid-Open Publication No. Hei1-125394] (19 mg) wasreacted by the general procedure according to Example 1 to obtain thetitle compound (15 mg, 46%) as white solid.

[0091]¹H-NMR (CD₃OD) δ: 3.02 (dd, 1H, J=4.9, 12.6 Hz, H-3eq), 1.92 (s,3H, NAc), 1.83 (t, 1H, J=12.1 Hz, H-3ax), 1.39-1.26 (m, 12H, 6CH₂), 0.90(t, 6H, J=6.6 Hz, 2CH₂CH₃).

Example 4 3-O-[Sodium{5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-β-D-galacto-2-nonulopyranosyl}onate]-1,2-di-O-hexyl-Sn-glycerol

[0092]3-O-[Sodium(5-acetamido-3,5-dideoxy-D-glycero-β-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-hexyl-Sn-glycerol[Japanese Patent Laid-Open Publication No. Hei1-125394] (26 mg) wasreacted by the general procedure according to Example 1 to obtain thetitle compound (22 mg, 48%) as white solid.

[0093]¹H-NMR (CD₃OD) δ: 2.83 (dd, 1H, J=5.1, 12.8 Hz, H-3eq), 2.03 (s,3H, NAc), 1.87 (t, 1H, J=12.3 Hz, H-3ax), 1.37-1.34 (m, 12H, 6CH₂), 0.90(t, 3H, J=6.8 Hz, CH₂CH₃), 0.90 (t, 3H, J=7.0 Hz, CH₂CH₃).

Example 5 3-O-[Sodium{5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosyl}onate]-1,2-di-O-decyl-Sn-glycerol

[0094]3-O-[Sodium(5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-decyl-Sn-glycerol[Japanese Patent Laid-Open Publication No. Hei1-125394] (190 mg) wasreacted by the general procedure according to Example 1 to obtain thetitle compound (268 mg, 88%) as white solid.

[0095]¹H-NMR (D₂O) δ: 2.94 (dd, 1H, J=5.0, 13.0 Hz, H-3eq), 1.99 (s, 3H,NAc), 1.96 (t, 1H, J=12.5 Hz, H-3ax), 1.44-1.21 (m, 28H, 14CH₂), 0.89(t, 6H, J=6.8 Hz, 2CH₂CH₃).

Example 6 3-O-[Sodium{5-acetamido-3,5-dideoxy4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-β-D-galacto-2-nonulopyranosyl}onate]-1,2-di-O-decyl-Sn-glycerol

[0096]3-O-[Sodium(5-acetamido-3,5-dideoxy-D-glycero-β-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-decyl-Sn-glycerol[Japanese Patent Laid-Open Publication No. Hei1-125394] (197 mg) wasreacted by the general procedure according to Example 1 to obtain thetitle compound (240 mg, 76%) as white solid.

[0097]¹H-NMR (D₂O) δ: 2.67 (dd, 1H, J=5.0, 13.0 Hz, H-3eq), 2.01 (s, 3H,NAc), 1.93 (t, 1H, J=12.2 Hz, H-3ax), 1.42-1.20 (m, 28H, 14CH₂), 0.88(t, 6H, J=6.8 Hz, 2CH₂CH₃).

Example 7 3-O-[Sodium{5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosyl}onate]-1,2-di-O-tetradecyl-Sn-glycerol

[0098]3-O-[Sodium(5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-tetradecyl-Sn-glycerol[Japanese Patent Laid-Open Publication No. Sho59-164798] (406 mg) wasreacted by the general procedure according to Example 1 to obtain thetitle compound (479 mg, 78%) as white solid.

[0099]¹H-NMR (CD₃OD) δ: 3.01 (dd, 1H, J=4.9, 11.9 Hz, H-3eq), 1.93 (s,3H, NAc), 1.74 (t, 1H, J=12.1 Hz, H-3ax), 1.38-1.25 (m, 44H, 22CH₂),0.90 (t, 6H, J=6.8 Hz, 2CH₂CH₃).

Example 8 3-O-[Sodium{5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycerol-β-D-galacto-2-nonulopyranosyl}ornate]-1,2-di-O-tetradecyl-Sn-glycerol

[0100]3-O-[Sodium(5-acetamido-3,5-dideoxy-D-glycero-βα-D-galacto-2-nonulopyranosyl)ornate]-1,2-di-O-tetradecyl-Sn-glycerol[Japanese Patent Laid-Open Publication No. Sho59-164798] (87 mg) wasreacted by the general procedure according to Example 1 to obtain thetitle compound (120 mg, 91%) as white solid.

[0101]¹H-MR (CD₃OD) δ: 2.83 (DD, 1H, J=5.5, 12.8 Hz, H-3eq), 2.03 (s,3H, ANC), 1.85 (t, 1H, J=12.0 Hz, H-ax), 1.45-1.20 (m, 44H, 22CH2), 0.90(t, 6H, J=7.0 Hz, 2CH2CH3).

Example 9 3-O-[Sodium{5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycerol-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-octadecyl-Sn-glycerol

[0102]3-O-[Sodium(5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-octadecyl-Sn-glycerol[Japanese Patent Laid-Open Publication No. Hei1-125394] (45 mg) wasreacted by the general procedure according to Example 1 to obtain thetitle compound (44 mg, 66%) as white solid.

[0103]¹H-NMR (CD₃OD) δ: 3.02 (dd, 1H, J=5.1, 13.2 Hz, H-3eq), 1.92 (s,3H, NAc), 1.82 (t, 1H, J=12.5 Hz, H-3ax), 1.38-1.22 (m, 60H, 30CH₂),0.90 (t, 6H, J=6.9 Hz, 2CH₂CH₃).

Example 10 3-O-[Sodium{5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosyl}onate]-2-O-(sodiumoxysulfonyl)-1-O-octadecyl-Sn-glycerol

[0104]3-O-[Sodium(5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1-O-octadecyl-Sn-glycerol[Japanese Patent Publication No. Hei1-125394] (132 mg) was reacted bythe general procedure according to Example 1 to obtain the titlecompound (125 mg, 53%) as white solid.

[0105]¹H-NMR (CD₃OD) δ: 3.07 (dd, 1H, J=5.1, 12.1 Hz, H-3eq), 1.95 (s,3H, NAc), 1.68 (t, 1H, J=12.1 Hz, H-3ax), 1.36-1.22 (m, 30H, 15CH₂),0.90 (t, 3H, J=7.0 Hz, CH₂CH₃).

Example 11 3-O-[Sodium{5-acetamido-3,5-dideoxy4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-β-D-galacto-2-nonulopyranosyl}onate]-2-O-(sodiumoxysulfonyl)-1-O-octadecyl-Sn-glycerol

[0106]3-O-[Sodium(5-acetamido-3,5-dideoxy-D-glycero-β-D-galacto-2-nonulopyranosyl)onate]-1-O-octadecyl-Sn-glycerol[Japanese Patent Publication No. Hei1-125394] (66 mg) was reacted by thegeneral procedure according to Example 1 to obtain the title compound(31 mg, 49%) as white solid.

[0107]¹H-NMR (CD₃OD) δ: 2.81 (dd, 1H, J=4.4, 12.1 Hz, H-3eq), 2.03 (s,3H, NAc), 1.88 (t, 1H, J=11.9 Hz, H-3ax), 1.38-1.17 (m, 30H, 15CH₂),0.90 (t, 3H, J=7.0 Hz, CH₂CH₃).

Example 12 3-O-[Sodium{5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosyl}onate]-1,2-di-O-docosyl-Sn-glycerol

[0108]3-O-[Sodium(5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-docosyl-Sn-glycerol[Japanese Patent Publication No. Hei1-125394] (502 mg) was reacted bythe general procedure according to Example 1 to obtain the titlecompound (447 mg, 54%) as white solid.

[0109]¹H-NMR (CD₃OD-D₂O, 1:1, 50° C.) δ: 2.96 (br. dd, 1H, H-3eq), 1.97(s, 3H, NAc), 1.83 (br. t, 1H, H-3ax), 1.40-1.21 (m, 76H, 38CH₂), 0.89(t, 6H, J=6.4 Hz, 2CH₂CH₃).

Example 13 3-O-[Sodium{5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-β-D-galacto-2-nonulopyranosyl}onate]-1,2-di-O-docosyl-Sn-glycerol

[0110]3-O-[Sodium(5-acetamido-3,5-dideoxy-D-glycero-β-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-docosyl-Sn-glycerol[Japanese Patent Publication No. Hei1-125394] (44 mg) was reacted by thegeneral procedure according to Example 1 to obtain the title compound(25 mg, 40%) as white solid.

[0111]¹H-NMR (CDCl₃-CD₃OD-D₂O, 3:4:2) δ: 2.75 (br. dd, 1H, H-3eq), 2.02(s, 3H, NAc), 1.89 (br. t, 1H, H-3ax), 1.40-1.18 (m, 76H, 38CH₂), 0.89(t, 6H, J=6.6 Hz, 2CH₂CH₃).

Example 14 3-S-[Sodium{5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosyl}onate]-1,2-di-O-tetradecyl-Sn-thioglycerol

[0112]3-S-[Sodium(5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-tetradecyl-Sn-thioglycerol[Japanese Patent Laid-Open Publication No. Sho64-52794] (41 mg) wasreacted by the general procedure according to Example 1 to obtain thetitle compound (47 mg, 77%) as white solid.

[0113]¹H-NMR (CD₃OD) δ: 3.17 (dd, 1H, J=5.1, 12.1 Hz, H-3eq), 1.92 (s,3H, NAc), 1.77 (t, 1H, J=11.7 Hz, H-3ax), 1.41-1.23 (m, 44H, 22CH₂),0.90 (t, 6H, J=7.1 Hz, 2CH₂CH₃).

Example 153-O-[Methyl(5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-eicosyl-Sn-glycerol

[0114] A 3-O-benzyl-1,2-di-O-eicosyl-Sn-glycerol:

[0115] 3-O-Benzyl-Sn-glycerol [Agric. Biol. Chem., 46, 255 (1982)] (300mg, 1.65 mmol), 1-bromo-eicosane (2.38 g, 6.58 mmol) and pulverizedsodium hydroxide (293 mg, 7.33 mmol) were azeotropically heated atreflux in benzene (10 ml) for 2 days to remove water from the mixture.After the reaction solution was diluted with ether and washed withwater, the organic layer was dried over anhydrous magnesium sulfate andthen condensed in vacuo. The residue was purified by silica gel columnchromatography (100 g of gel, hexane:toluene=3:2) to obtain the titlecompound (929 mg, 76%) as white powder.

[0116]¹H-NMR (CDCl₃) δ: 7.34-7.26 (m, 5H, C₆H₅), 4.55 (s, 2H, CH₂Ph),1.25 (m, 68H, 34H₂), 0.88 (t, 6H, J=6.6 Hz, 2CH₂CH₃).

[0117] B 1,2-Di-O-eicosyl-Sn-glycerol:

[0118] 3-O-benzyl-1,2-di-O-eicosyl-Sn-glycerol (1.47 g, 1.98 mmol) and10% palladium-charcoal (200 mg). were stirred in ethyl acetate (30 ml)under the hydrogen atmosphere at room temperature for 18 h. The reactionmixture was filtered through celite, and the filtrate was concentratedin vacuo to obtain the title compound (934 mg, 72%) as white powder.

[0119]¹H-NMR (CDCl₃) δ: 1.25 (m, 68H, 34CH₂), 0.88 (t, 6H, J=6.6 Hz,2CH₂CH₃).

[0120] C3-O-[Methyl(5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-eisosyl-Sn-glycerol:

[0121] A mixture comprising 1,2-di-O-eicosyl-Sn-glycerol (497 mg, 0.76mmol), methyl5-acetamido-4,7,8,9-tetra-O-acetyl-2-chloro-2,3,5-trideoxy-D-glycero-β-D-galacto-2-nonulopyranosonate[Chem. Ber., 99, 611 (1966)] (422 mg, 0.83 mmol), mercury (II) cyanide(336 mg, 1.33 mmol), mercury (II) bromide (478 mg, 1.33 mmol) and driedmolecular sieve 4A (1.0 g) in anhydrous chloroform (5.0 ml) was stirredunder the nitrogen atmosphere at room temperature overnight. After thereaction mixture was filtered, the filtrate was concentrated in vacuo,and purified by intermediary pressure silica gel column chromatography(125 g of gel, toluene:ethyl acetate=3:2) to obtain the title compound(390 mg, 46%) as white power.

[0122]¹H-NMR (CDCl₃) δ: 3.79 (s, 3H, OCH₃), 2.60 (dd, 1H, J=4.8, 12.8Hz, H-3eq.), 2.14, 2.13, 2.06, 2.04, 1.88 (5s, 15H, 5Ac), 1.98 (t, 1H,J=12.8 Hz, H-3ax.), 1.25 (m, 68H, 34CH₂), 0.88 (t, 6H, J=7.0 Hz,2CH₂CH₃).

Example 163-O-[(5-Acetamido-3,5-dideoxy-D-glycero-α-D-galacto-nonulopyranosonicacid)-2-yl]-1,2-di-O-eicosyl-Sn-glycerol

[0123]3-O-[Methyl(5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-eicosyl-Sn-glycerol(455 mg, 0.40 mmol) and sodium methoxide was stirred in a mixture (13ml) of THF:methanol (1:1) overnight. To this reaction mixture was added3N sodium hydroxide (380 μl), and the mixture was stirred at 60° C. for1 h. After the reaction mixture was concentrated in vcuo, the residuewas dissolved in ethanol and water, and the solution was adjusted to pH2 with formic acid. Precipitates were collected by filtration, washedsuccessively with water, methanol and ether, and dried in vacuo toobtain the title compound (365 mg, 96%) as white powder.

[0124]¹H-NMR (CDCl₃-CD₃OD, 1:1) δ: 2.79 (dd, 1H, J=3.9, 12.3 Hz,H-3eq.), 2.03 (s, 3H, NAc), 1.69 (t, 1H, J=11.5 Hz, H-3ax.), 1.43-1.20(m, 68H, 34CH₂), 0.89 (t, 6H, J=7.0 Hz, 2CH₂CH₃).

Example 17 3-O-[Sodium{5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-eicosyl-Sn-glycerol

[0125]3-O-[(5-Acetamido-3,5-dideoxy-D-glycero-α-D-galacto-nonulopyranosonicacid)-2-yl]-1,2-di-O-eicosyl-Sn-glycerol (241 mg) was reacted by thegeneral procedure according to Example 1 to obtain the title compound(236 mg, 67%) as white solid.

[0126]¹H-NMR (CD₃OD) δ: 3.02 (dd, 1H, J=4.4, 12.4 Hz, H-3eq.), 1.92 (s,3H, NAc), 1.80 (t, 1H, J=11.0 Hz, H-3ax.), 1.44-1.13 (m, 68H, 34CH₂),0.909 (t, 6H, J=6.8 Hz, 2CH₂CH₃).

Example 183-O-[Methyl(5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl}onate]-1,2-di-O-tetracosyl-Sn-glycerol

[0127] A 3-O-Benzyl-1,2-di-O-tetracosyl-Sn-glycerol:

[0128] 3-O-Benzyl-Sn-glycerol [Agric. Biol. Chem., 46, 255 (1982)] and1-bromotetracosane [J. Am. Chem. Soc., 115, 3840 (1993)] were reacted bythe general procedure according to Example 1 to obtain the titlecompound (236 mg, 67%) as white solid.

[0129]¹H-NMR (CDCl₃) δ: 7.34-7.26 (m, 5H, C₆H₅), 4.55 (s, 2H, CH₂Ph),1.25 (m, 84H, 42CH₂), 0.88 (t, 6H, J=6.6 Hz, 2CH₂CH₃).

[0130] B 1,2-Di-O-tetracosyl-Sn-glycerol:

[0131] 3-O-Benzyl-1,2-di-O-tetracosyl-Sn-glycerol (1.47 g) was reactedin a mixture (30 ml) of toluene-ethyl acetate (1:1) by the generalprocedure according to 15-B to obtain the title compound (0.80 g, 66%)as white solid.

[0132]¹H-NMR (CDCl₃) δ: 1.25 (m, 84H, 42CH₂), 0.88 (t, 6H, J=6.6 Hz,2CH₂CH₃).

[0133] C3-O-[Methyl(5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl}onate]-1,2-di-O-tetracosyl-Sn-glycerol:

[0134] 1,2-Di-O-tetracosyl-Sn-glycerol (497 mg, 0.76 mmol) and methyl5-acetamido-4,7,8,9-tetra-O-acetyl-2-chloro-2,3,5-trideoxy-D-glycero-β-D-galacto-2-nonulopyranosonate[Chem. Ber., 99, 611 (1966)] (422 mg, 0.83 mmol) were reacted by thegeneral procedure according to 15-C to obtain the title compound (224mg, 14%) as white solid.

[0135]¹H-NMR (CDCl₃) δ: 3.79 (s, 3H, OCH₃), 2.60 (dd, 1H, J=4.8, 12.8Hz, H-3eq), 2.14, 2.13, 2.04, 2.03, 1.88 (5s, 15H, 5Ac), 1.98 (t, 1H,J=12.8 Hz, H-3ax), 1.25 (m, 84H, 42CH₂), 0.88 (t, 6H, J=6.6 Hz,2CH₂CH₃).

Example 193-O-[(5-Acetamido-3,5-dideoxy-D-glycero-α-D-galacto-nonulopyranosonicacid)-2-yl]-1,2-di-O-tetracosyl-Sn-glycerol

[0136]3-O-[Methyl(5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl}onate]-1,2-di-O-tetracosyl-Sn-glycerol(240 mg) was reacted by the general procedure according to Example 16 toobtain the title compound (160 mg, 79%) as white solid.

[0137]¹H-NMR (CDCl₃-CD₃OD, 1:1) δ: 2.81 (dd, 1H, J=3.7, 12.1 Hz, H-3eq),2.03 (s, 3H, NAc), 1.66 (t, 1H, J=11.3 Hz, H-3ax), 1.38-1.16 (m, 84H,42CH₂), 0.89 (t, 6H, J=6.8 Hz, 2CH₂CH₃).

Example 20 3-O-[Sodium{5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosyl}onate]-1,2-di-O-tetracosyl-Sn-glycerol

[0138]3-O-[(5-Acetamido-3,5-dideoxy-D-glycero-α-D-galacto-nonulopyranosonicacid) reacted by the general procedure according to Example 1 to obtainthe title compound (21 mg, 19%) as white solid.

[0139]¹H-NMR (CD₃OD-D₂O, 1:1, 50° C.) δ: 2.96 (br. dd, 1H, H-3eq), 2.01(s, 3H, NAc), 1.81 (br. t, 1H, H-3ax), 1.36-1.10 (m, 84H, 42CH₂), 0.89(t, 6H, J=6.4 Hz, 2CH₂CH₃).

Example 213-O-[Methyl(5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-hexacosyl-Sn-glycerol

[0140] A 3-O-Benzyl-1,2-di-O-hexacosyl-Sn-glycerol:

[0141] 3-O-Benzyl-Sn-glycerol [Agric. Biol. Chem., 40, 391 (1976)] (242mg) and 1-bromohexacosane [Agric. Biol. Chem., 46, 255 (1982)] (1.30 g)were reacted by the general procedure according to Example 15-A. Then,the compound thus obtained was, without purification, subjected to thedebenzylation by the general procedure according to Example 15-B toobtain the title compound (615 mg, 57%) as white solid.

[0142]¹H-NMR (CDCl₃) δ: 1.25 (m, 92H, 46CH₂), 0.88 (t, 6H, J=6.6 Hz,2CH₂CH₃).

[0143] B3-O-[Methyl(5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-hexacosyl-Sn-glycerol:

[0144] 1,2-Di-O-hexacosyl-Sn-glycerol (610 mg, 0.74 mmol) and methyl5-acetamido-4,7,8,9-tetra-O-acetyl-2-chloro-2,3,5-trideoxy-D-glycero-β-D-galacto-2-nonulopyranosonate[Chem. Ber., 99, 611 (1966)] (454 mg, 0.89 mmol) were reacted by thegeneral procedure according to Example 15-C to obtain the title compound(76 mg, 8%) as white solid.

[0145]¹H-NMR (CDCl₃) δ: 3.79 (s, 3H, OCH₃), 2.60 (dd, 1H, J=4.8, 12.8Hz, H-3eq), 2.14, 2.13, 2.04, 2.02, 1.88 (5s, 15H, 5Ac), 1.97 (t, 1H,J=12.8 Hz, H-3ax), 1.25 (m, 92H, 46CH₂), 0.88 (t, 6H, J=7.0 Hz,2CH₂CH₃).

Example 223-O-[(5-Acetamido-3,5-dideoxy-D-glycero-α-D-galacto-nonulopyranosonicacid)-2-yl]-1,2-di-O-hexacosyl-Sn-glycerol

[0146]3-O-[Methyl(5-acetamido4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-hexacosyl-Sn-glycerol(76 mg) was reacted by the general procedure according to Example 16 toobtain the title compound (51 mg, 78%) as white solid.

[0147]¹H-NMR (CDCl₃-CD₃OD, 1:1, 60° C.) δ: 2.76 (dd, 1H, J=4.4, 12.8 Hz,H-3eq), 2.02 (s, 3H, NAc), 1.75 (t, 1H, J=11.7 Hz, H-3ax), 1.42-1.14 (m,92H, 46CH₂), 0.89 (t, 6H, J=6.8 Hz, 2CH₂CH₃).

Example 23 3-O-[Sodium{5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosyl}onate]-1,2-di-O-hexacosyl-Sn-glycerol

[0148]3-O-[(5-Acetamido-3,5-dideoxy-D-glycero-α-D-galacto-nonulopyranosonicacid)-2-yl]-1,2-di-O-hexacosyl-Sn-glycerol (47 mg) was reacted by thegeneral procedure according to Example 1 to obtain the title compound(29 mg, 45%) as white solid.

[0149]¹H-NMR (CD₃OD-D₂O, 1:1, 60° C.) δ: 2.96 (br. dd, 1H, H-3eq), 1.96(s, 3H, NAc), 1.79 (br. t, 1H, H-3ax), 1.42-1.14 (m, 92H, 46CH₂), 0.88(br. t, 6H, J=6.2 Hz, 2CH₂CH₃).

Example 24 3β-[Sodium{5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosyl}onate]-5-cholestene

[0150]3β-[(5-Acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosonicacid)-2-yl]-5-cholestene [Japanese Patent Laid-Open Publication No. Sho61-243096] (387 mg) was reacted by the general procedure according toExample 1 to obtain the title compound (232 mg, 37%) as white solid.

[0151]¹H-NMR (CD₃OD-D₂O, 1:1) δ: 2.92 (dd, 1H, J=4.5, 11.5 Hz, H-3eq),1.96 (s, 3H, NAc), 0.99 (s, 3H, CH₃-19 chole), 0.68 (s, 3H, CH₃-18chole).

Example 25 Sodium[oleyl 5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosid]onate:

[0152] A Sodium(oleyl5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid)onate:

[0153] To a solution of methyl(oleyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid)onate[Japanese Patent Laid-Open Publication No. Sho 63-264493] (115 mg, 0.154mmol) dissolved in methanol (1.5 ml) was added 3N sodium hydroxide (0.30ml), and the mixture was stirred at room temperature for 16 h. Thereaction solution was evaporated to dryness in vcuo, and the residue waspurified by gel chromatography (Sephadex LH-20, 180 ml, methanol) toobtain the title compound (78 mg, 87%) as white solid.

[0154]¹H-NMR (CD₃OD) δ: 5.34 (m, 2H, CH═CH), 2.81 (dd, 1H, J=4.0, 12.1Hz, H-3eq), 2.00 (s, 3H, NAc), 1.59 (t, 1H, J=11.7 Hz, H-3ax), 1.42-1.23(m, 20H, 10CH₂), 0.90 (t, 3H, J=7.0 Hz, CH₂CH₃).

[0155] B Sodium[oleyl 5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosid]onate:

[0156] Sodium(oleyl5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid)onate(62 mg) was reacted by the general procedure according to Example 1 toobtain the title compound (86 mg, 82%) as white solid.

[0157]¹H-NMR (CD₃OD) δ: 3.03 (dd, 1H, J=3.7, 12.3 Hz, H-3eq), 2.19 (m,4H, CH₂CH═CHCH₂), 1.92 (s, 3H, NAc), 1.73 (t, 1H, J=12.1 Hz, H-3ax),1.42-1.22 (m, 20H, 10CH₂), 0.90 (t, 3H, J=6.8 Hz, CH₂CH₃).

Example 26 Sodium[octadecyl5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosid)onate

[0158] A Methyl(octadecyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid)onate:

[0159] To a solution of methyl(oleyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid)onate[Japanese Patent Laid-Open Publication No. Sho 63-264493] (111 mg, 0.149mmol) dissolved in ethanol (1.5 ml) was added 10% palladium-charcoal (16mg) was added, and the mixture was stirred under the hydrogen atmosphereat room temperature for 18 h. The reaction solution was filtered throughcelite to obtain the title compound (107 mg, 96%) as white solid.

[0160]¹H-NMR (CDCl₃) δ: 3.79 (s, 3H, COOCH₃), 2.58 (dd, 1H, J=4.8, 12.8Hz, H-3eq), 2.15, 2.14, 2.04, 2.03, 1.88 (5s, 15H, 5Ac), 1.95 (t, 1H,J=12.6 Hz, H-3ax), 1.38-1.19 (m, 30H, 15CH₂), 0.88 (t, 3H, J=7.0 Hz,CH₂CH₃).

[0161] B Sodium(octadecyl5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid)onate:

[0162] Methyl(octadecyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid)onate(107 mg) was reacted by the general procedure according to Example 25-Ato obtain the titled compound (70 mg, 83%) as white solid.

[0163]¹H-NMR (CD₃OD) δ: 2.81 (dd, 1H, J=3.8, 12.3 Hz, H-3eq), 2.00 (s,3H, NAc), 1.59 (t, 1H, J=11.9 Hz, H-3ax), 1.38-1.22 (m, 30H, 15CH₂),0.90 (t, 3H, J=6.8 Hz, CH₂CH₃).

[0164] C Sodium[octadecyl5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosid]onate:

[0165] Sodium(octadecyl5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid)onate(72 mg) was reacted by the general procedure according to Example 1 toobtain the title compound (109 mg, 92%) as white solid.

[0166]¹H-NMR (CD₃OD) δ: 3.02 (dd, 1H, J=3.7, 11.4 Hz, H-3eq), 1.92 (s,3H, NAc), 1.73 (t, 1H, J=11.7 Hz, H-3ax), 1.40-1.31 (m, 30H, 15CH₂),0.90 (t, 3H, J=7.0 Hz, CH₂CH₃).

Example 27 Methyl[2,2-bis(docosyloxymethyl)propyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid]onate

[0167] A 2,2- bis(docosyloxymethyl)propanol:

[0168] 1,1,1-tris(hydroxymethyl)ethane (1.0 g, 8.32 mmol) and sodiumhydride (732 mg, 18.3 mmol) were stirred in dehydrated dimethylformamide(30 ml) at room temperature for 15 min. Then, the reaction solution wascooled in ice, and docosyl bromide (7.1 g, 18.3 mmol) and benzene (10ml) were added thereto, and the resulting mixture was stirred at roomtemperature for 16 h. The reaction solution was concentrated in vcuo,and the residue was suspended in chloroform, and washed with 2N HCl. Theorganic layer was dried over anhydrous magnesium sulfate, andconcentrated in vacuo. The residue was purified by silica gel columnchromatography (130 g of gel, toluene/ethyl acetate, 19:1) to obtain thetitle compound (2.32 g, 38%) as white powder.

[0169]¹H-NMR (CDCl₃) δ: 3.56 (d, 2H, J=5.9 Hz, CH₂OH), 1.26 (m, 76H,38CH₂), 0.88 (t, 6H, J=6.6 Hz, 2CH₂CH₃), 0.85 (s, 3H, CCH₃).

[0170] B Methyl[2,2-bis(docosyloxymethyl)propyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid]onate:

[0171] 2,2-Bis(docosyloxymethyl)propanol (1.20 g, 1.63 mmol) and methyl5-acetamido-4,7,8,9-tetra-O-acetyl-2-chloro-2,3,5-trideoxy-D-glycero-β-D-galacto-2-nonulopyranosonate[Chem. Ber., 99, 611 (1966)] (913 mg, 1.79 mmol) were reacted by thegeneral procedure according to Example 15-C to obtain the title compound(786 mg, 40%) as white solid.

[0172]¹H-NMR (CDCl₃) δ: 3.78 (s, 3H, COOCH₃), 2.57 (dd, 1H, J=4.4, 12.8Hz, H-3eq.), 2.13, 2.12, 2.04, 2.02, 1.88 (5s, 15H, 5Ac), 1.95 (t, 1H,J=12.5 Hz, H-3ax.), 1.25 (m, 76H, 38CH₂), 0.90 (s, 3H, CCH₃), 0.88 (t,6H, J=7.0 Hz, 2CH₂CH₃).

Example 28 2,2-Bis(docosyloxymethyl)propyl5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosidonic acid

[0173] Methyl[2,2-bis(docosyloxymethyl)propyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid]onate(786 mg) was reacted by the general procedure according to Example 16 toobtain the title compound (599 mg, 90%) as white solid.

[0174]¹H-NMR (CDCl₃-CD₃OD, 1:1, 60° C.) δ: 2.79 (dd, 1H, J=4.8, 12.8 Hz,H-3eq), 2.02 (s, 3H, NAc), 1.75 (t, 1H, J=11.9 Hz, H-3ax), 1.36-1.22 (m,76H, 38CH₂), 0.92 (s, 3H, CCH₃), 0.89 (t, 6H, J=6.8 Hz, 2CH₂CH₃).

Example 29 Sodium[2,2-bis(docosyloxymethyl)propyl5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosid]onate

[0175] 2,2-Bis(docosyloxymethyl)propyl5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosidonic acid(94 mg) was reacted by the general procedure according to Example 1 toobtain the title compound (63 mg, 47%) as white solid. (599 mg, 90%) aswhite solid.

[0176]¹H-NMR (CD₃OD-D₂O, 1:1, 60° C.) δ: 2.93 (br. dd, 1H, H-3eq), 1.96(s, 3H, NAc), 1.80 (br. t, 1H, H-3ax), 1.43-1.16 (m, 76H, 38CH₂), 0.92(s, 3H, CCH₃), 0.89 (t, 6H, J=6.6 Hz, 2CH₂CH₃).

Example 30 Methyl[2,2-bis(eicosyloxymethyl)propyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid]onate

[0177] A 2,2-Bis(eicosyloxymethyl)propanol:

[0178] 1,1,1-tris(hydroxymethyl)ethane (0.50 g, 4.16 mmol) and eicosylbromide (3.30 g, 9.15 mmol) were reacted by the general procedureaccording to Example 27-A to obtain the title compound (1.43 g, 51%) aswhite solid.

[0179]¹H-NMR (CDCl₃) δ: 3.57 (d, 2H, J=5.9 Hz, CH₂OH), 1.26 (m, 68H,34CH₂), 0.88 (t, 6H, J=7.0 Hz, 2CH₂CH₃), 0.85 (s, 3H, CCH₃).

[0180] B Methyl[2,2-bis(eicosyloxymethyl)propyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid]onate:

[0181] 2,2-bis(eicosyloxymethyl)propanol (267 mg, 0.392 mmol) and methyl5-acetamido-4,7,8,9-tetra-O-acetyl-2-chloro-2,3,5-trideoxy-D-glycero-β-D-galacto-2-nonulopyranosonate[Chem. Ber., 99, 611 (1966)] (200 mg, 0.392 mmol) were reacted by thegeneral procedure according to Example 15-C to obtain the title compound(175 mg, 39%) as white solid.

[0182]¹H-NMR (CDCl₃) δ: 3.78 (s, 3H, COOCH₃), 2.58 (dd, 1H, J=4.8, 12.8Hz, H-3eq.), 2.13, 2.13, 2.04, 2.02, 1.89 (5s, 15H, 5Ac), 1.95 (dd, 1H,J=12.5, 12.8 Hz, H-3ax.), 1.25 (m, 68H, 34CH₂), 0.90 (s, 3H, CCH₃), 0.88(t, 6H, J=7.0 Hz, 2CH₂CH₃).

Example 31 2,2-bis(eicosyloxymethyl)propyl5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosidonic acid

[0183] Methyl[2,2-bis(eicosyloxymethyl)propyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid]onate(174 mg) was reacted by the general procedure according to Example 16 toobtain the title compound (127 mg, 87%) as white solid.

[0184]¹H-NMR (CDCl₃-CD₃OD, 1:1) δ: 2.75 (dd, 1H, J=4.6, 12.6 Hz, H-3eq),2.03 (s, 3H, NAc), 1.75 (t, 1H, J=11.7 Hz, H-3ax), 1.43-1.19 (m, 68H,34CH₂), 0.92 (s, 3H, CCH₃), 0.89 (t, 6H, J=7.0 Hz, 2CH₂CH₃).

Example 32 Sodium[2,2-bis(eicosyloxymethyl)propyl5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosid)onate

[0185] 2,2-bis(eicosyloxymethyl)propyl5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosidonic acid(103 mg) acid was reacted by the general procedure according to Example1 to obtain the title compound (96 mg, 64%) as white solid.

[0186]¹H-NMR (CD₃OD-D₂O, 1:1, 60° C.) δ: 2.96 (br. dd, 1H, H-3eq), 1.98(s, 3H, NAc), 1.78 (t, 1H, J=12.6 Hz, H-3ax), 1.37-1.17 (m, 68H, 34CH₂),0.94 (s, 3H, CCH₃), 0.90 (t, 6H, J=6.8 Hz, 2CH₂CH₃).

Example 33 Methyl(3,5-didocosyloxyphenyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid)onate

[0187] A 3,5-didocosyloxyphenol:

[0188] After phloroglucinol (2.0 g, 15.9 mmol) and sodium hydride (1.59g, 39.7 mmol) were stirred in dehydrated dimethylformamide (30 ml) atroom temerature for 15 min, docosyl bromide (13.0 mg, 33.3 mmol) andbenzene (30 ml) were added thereto, and the resulting mixture wasstirred at 40° C. for 2 days. The reaction solution was diluted withchloroform, washed with 2N hydrochloric acid, and the organic layer wasdried over anhydrous magnesium sulfate. The solvent was concentrated invacuo, and the residue was purified by silica gel column chromatography(90 g of the gel, hexane/ethyl acetate, 6:1) to obtain the titlecompound (2.40 g, 20%) as light yellow powder.

[0189]¹H-NMR (CDCl₃) δ: 6.06 (d, 1H, J=2.2 Hz, H-4 phenyl), 5.99 (2d,2H, J=2.2 Hz, H-2,6 phenyl), 3.89 (t, 4H, J=6.8 Hz, 2OCH₂), 1.39-1.17(m, 72H, 36CH₂), 0.88 (t, 6H, J=6.8 Hz, 2CH₂CH₃).

[0190] B Methyl(3,5-didocosyloxyphenyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid)onate:

[0191] 3,5-Didocosyloxyphenol (1.20 g, 1.61 mmol) and sodium hydride(77.5 g, 1.94 mmol) were stirred in dimethylformamide:benzene (1:1, 40ml) at room temperature for 20 min. Then, to this mixture was addedmethyl5-acetamido-4,7,8,9-tetra-O-acetyl-2-chloro-2,3,5-trideoxy-D-glycero-β-D-galacto-2-nonulopyranosonate[Chem. Ber., 99, 611 (1966)] (823 mg, 1.61 mmol), and the resultingmixture was stirred at room temperature for 3 h. After the reactionsolution was diluted with chloroform, and washed with 2N-hydrochloricacid and, the organic layer was dried over anhydrous magnesium sulfate.The solvent was condensed in vacuo, and the residue was purified bysilica gel column chromatography (90 g of the gel, toluene/ethylacetate, 2:3) to obtain the title compound (386 mg, 20%) light yellowpowder.

[0192]¹H-NMR (CDCl₃) δ: 6.28 (2d, 2H, J=1.8 Hz, H-2,6phenyl), 6.22 (d,1H, J=1.8 Hz, H-4phenyl), 3.78 (s, 3H, COOCH₃), 2.58 (dd, 1H, J=4.8,12.8 Hz, H-3eq.), 2.14, 2.10, 2.03, 2.01, 1.90 (5s, 15H, 5Ac), 1.26 (m,76H, 38CH₂), 0.88 (t, 6H, J=6.6 Hz, 2CH₂CH₃).

Example 34 3,5-Didocosyloxyphenyl5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyrasidonic acid

[0193] Methyl(3,5-didocosyloxyphenyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid)onate(385 mg) was reacted by the general procedure according to Example 16 toobtain the title compound (163 mg, 50%) as light yellow solid.

[0194]¹H-NMR (CDCl₃-CD₃OD, 1:1, 60° C.) δ: 6.42 (2d, 2H, J=2.2 Hz,H-2,6phenyl), 6.20 (d, 1H, J=2.2 Hz, H-4phenyl), 2.86 (dd, 1H, J=4.2,12.6 Hz, H-3eq), 2.02 (s, 3H, NAc), 1.95 (t, 1H, J=11.9 Hz, H-3ax),1.40-1.23 (m, 72H, 36CH₂), 0.89 (t, 6H, J=7.0 Hz, 2CH₂CH₃).

Example 35Sodium[3,5-Didocosyloxyphenyl-5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-β-D-galacto-2-nonulopyranosid)onate

[0195] 3,5-Didocosyloxyphenyl5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosidonic acid(16 mg) was reacted by the general procedure according to Example 1 toobtain the title compound (11 mg, 47%) as white solid.

[0196]¹H-NMR (CD₃OD-D₂O, 1:1, 50° C.) δ: 6.42 (br. s, 2H, H-2,6phenyl),6.17 (br. s, 1H, H-4phenyl), 2.93 (br. dd, 1H, H-3eq), 2.00 (s, 3H,NAc), 1.38-1.18 (m, 72H, 36CH₂), 0.90 (t, 6H, J=7.0 Hz, 2CH₂CH₃).

Example 363-O-[Methyl(5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-docosanoyl-Sn-glycerol

[0197] A 3-O-Benzyl-1,2-di-O-docosanoyl-Sn-glycerol:

[0198] 3-O-Benzyl-Sn-glycerol [Agric. Biol. Chem., 40, 391 (1976)] (0.67g, 3.68 mmol), behenic acid (3.76 g, 11.04 mmol),dicyclohexylcarbodiimide (2.28 g, 11.04 mmol) and4-dimethylaminopyridine (0.13 g, 1.06 mmol) were stirred in pyridine (37ml)at room temperature for 22 h. The reaction solution was filtered, andevaporated to dryness in vacuo. The residue was purified by silica gelcolumn chromatography (using 90 g of gel, hexane/ethyl acetate, 9:1) toobtain the title compound (1.55 g, 51%) as light yellow powder.

[0199]¹H-NMR (CDCl₃) δ: 7.30 (m, 5H, C₆H₅), 5.24 (m, 1H, H-2), 4.56 (d,1H, J=12.1 Hz, C₆H₅CH), 4.52 (d, 1H, J=12.1 Hz, C₆H₅CH), 1.25 (m, 72H,36CH₂), 0.88 (t, 6H, J=7.0 Hz, 2CH₂CH₃).

[0200] B 1,2-Di-O-docosanoyl-Sn-glycerol:

[0201] 3-O-Benzyl-1,2-di-O-docosanoyl-Sn-glycerol (1.25 g) was reactedby the general procedure according to Example 15-B to obtain the titlecompound (0.98 g, 88%) as white solid.

[0202]¹H-NMR (CDCl₃) δ: 5.08 (m, 1H, H-2), 4.32 (dd, 1H, J=4.6, 11.9 Hz,H-3), 4.24 (dd, 1H, J=5.7, 11.9 Hz, H-3), 3.73 (m, 2H, CH₂-1), 1.25 (m,72H, 36CH₂), 0.88 (t, 6H, J=7.0 Hz, 2CH₂CH₃).

[0203] C3-O-[Methyl(5-acetamido-4,7,8,9-tetra-O-levulinoyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-docosanoyl-Sn-glycerol:

[0204] Methyl5-acetamido-3,5-dideoxy-D-glycero-D-galacto-2-nonulopyranosonate [Chem.Ber., 99, 611 (1966)] (3.23 g, 10.0 mmol), 4-dimethylaminopyridine (0.61g, 0.50 mmol), levulinic acid (12.3 ml, 120 mmol) anddicyclohexylcarbodiimide (24.76 g, 120 mmol) were stirred in pyridine(30 ml) at room temperature for two days. The reaction solution wasfiltered through celite, and the filtrate was evaporated to dryness invacuo. The residue was purified by silica gel column chromatography(with 170 g of gel, chloroform/methanol, 24:1) to obtain thepenta-levulinoyl derivative (4.62 g, 65%) of the title compound. Thenthe penta-levulinoyl derivative (2.50 g, 3.07 mmol) was dissolved inacetyl chloride (30 ml), saturated with hydrogen chloride gas at 0° C.,left at standing at 2° C. for five days. The reaction solution wasevaporated to dryness in vacuo to obtain the chloride derivative (2.15g, 95%) of the title compound. Then, the chloride derivative (1.22 g,1.66 mmol) and 1,2-di-O-docosanoyl-Sn-glycerol (0.89 g, 1.21 mmol) werereacted by the general procedure according to Example 15-C to obtain thetitle compound (64 mg, 4%) as white solid.

[0205]¹H-NMR (CDCl₃) δ: 5.06 (dt, 1H, J=4.6, 11.7 Hz, H-4), 3.81 (s, 3H,COOCH₃), 2.19 (6H), 2.18, 2.17 (3s, 12H, 3CH₃CO), 1.89 (s, 3H, NAc),1.85 (t, 1H, J=12.5 Hz, H-3ax), 1.25 (m, 72H, 36CH₂), 0.88 (t, 6H, J=7.0Hz, 2CH₂CH₃).

Example 373-O-[(5-Acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosonicacid-2-yl]-1,2-di-O-docosanoyl-Sn-glycerol

[0206]3-O-[Methyl(5-acetamido-4,7,8,9-tetra-O-levulinoyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-docosanoyl-Sn-glycerol(52 mg, 0.036 mmol) and anhydrous lithium iodide (49 mg, 0.366 mmol)were stirred in dehydrated pyridine (1.8 ml) at 80° C. for 6 h. Thereaction solution was directly purified by gel chromatography (usingSephadex LH-20, 100 ml, chloroform/methanol, 1:1) to obtain lithium saltof title compound (34.6 mg, 67%) as white solid. The lithium salt (34.6mg, 0.0242 mmol) was dissolved in methanol (0.5 ml) and chloroform (0.5ml), added with hydrazine acetate (48 mg, 0.524 mmol), and the mixturewas stirred at room temperature for 10 min. The reaction solution wasadjusted to pH 4 with 0.1N-hydrochloric acid, and purified by gelchromatography (Sephadex LH-20, 75 ml, chloroform/methanol, 1:1) toobtain the title compound (23 mg, 92%) as white solid.

[0207]¹H-NMR (CDCl₃-CD₃OD, 1:1, 40° C.) δ: 2.76 (dd, 1H, J=4.2, 12.6 Hz,H-3eq), 2.03 (s, 3H, NAc), 1.74 (t, 1H, J=11.9 Hz, H-3ax), 1.28 (m, 72H,36CH₂), 0.89 (t, 6H, J=7.0 Hz, 2CH₂CH₃).

Example 38 3-O-[Sodium{5-acetamido-3,5-dideoxy-4,7,8,9-tretra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosyl}onate]-1,2-di-O-docosanoyl-Sn-glycerol

[0208]3-O-[(5-Acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosonicacid)-2-yd]-1,2-di-O-docosanoyl-Sn-glycerol (12 mg) was reacted by thegeneral procedure according to Example 1 to obtain the title compound (9mg, 53%) as white solid.

[0209]¹H-MR (CD3OD-D2O, 1:1, 50° C.) δ: 2.94 (BR. DD, 1H, H-3eq), 1.95(s, 3H, ANC), 1.71 (t, 1H, J=11.0 Hz, H-ax), 1.26 (m, 72H, 36CH2), 0.87(t, 6H, J=6.8 Hz, 2CH2CH3).

Example 393-O-[Methyl(4,5,7,8,9-penta-O-acetyl-3-deoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-docosyl-Sn-glycerol

[0210] 1,2-Di-O-docosyl-Sn-glycerol [Japanese Patent Laid-OpenPublication No. Hei 1-125394] (467 mg) and methyl4,5,7,8,9-penta-O-acetyl-2-chloro-2,3-dideoxy-D-glycero-β-D-galacto-2-nonulopyranosonate[Chem. Pharm. Bull., 39, 3140 (1991)] (281 mg) were reacted by thegeneral procedure according to Example 15-C to obtain the title compound(313 mg, 48%) as white solid.

[0211]¹H-NMR (CDCl₃) δ: 3.80 (s, 3H, OCH₃, 2.68 (dd, 1H, J=4.0, 12.8 Hz,H-3eq), 2.15, 2.09, 2.04, 2.00, 2.00 (5s, 15H, 5Ac), 1.25 (m, 76H,38CH₂), 0.88 (t, 6H, J=6.6 Hz, 2CH₂CH₃).

Example 40 3-O-[(3-Deoxy-D-glycero-α-D-galacto-2-nonulopyranosonic acidacid)-2-yl]-1,2-di-O-docosyl-Sn-glycerol

[0212]3-O-[Methyl(4,5,7,8,9-penta-O-acetyl-3-deoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-docosyl-Sn-glycerol(313 mg) was reacted by the general procedure according to Example 16 toobtain the title compound (186 mg, 75%) as white solid.

[0213]¹H-NMR (CDCl₃-CD₃OD, 1:1, 60° C.) δ: 2.68 (dd, 1H, J=4.6, 12.6 Hz,H-3eq), 1.73 (t, 1H, J=12.1 Hz, H-3ax), 1.29 (m, 76H, 38CH₂), 0.89 (t,6H, J=7.0 Hz, 2CH₂CH₃).

Example 41 3-O-[Sodium{3-deoxy-4,5,7,8,9-penta-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosyl}onate]-1,2-di-O-docosyl-Sn-glycerol

[0214] 3-O-[(3-Deoxy-D-glycero-α-D-galacto-2-nonulopyranosonicacid)-2-yl]-1,2-di-O-docosyl-Sn-glycerol (97 mg) was reacted by thegeneral procedure according to Example 1 to obtain the title compound(15 mg, 10%) as white solid.

[0215]¹H-NMR (CD₃OD-D₂O, 1:1, 50° C.) δ: 2.89 (br. dd, 1H, H-3eq), 1.81(br. t, 1H, H-3ax), 1.26 (m, 76H, 38CH₂), 0.86 (t, 6H, J=7.0 Hz,2CH₂CH₃).

Example 421-O-[Methyl(5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-L-2,3-di-O-docosyl-Sn-glycerol

[0216] A L-1-O-Benyl-2,3-di-O-isopropylidene-Sn-glycerol:

[0217] After L-2,3-di-O-isopropylidene-Sn-glycerol (5.00 g, 37.8 mmol)and 60% sodium hydride (3.03 g, 75.8 mmol) were stirred in dehydrateddimethylformamide (110 ml) at room temperature for 10 min, benzylbromide (6.7 ml, 56.7 mmol) was added, and the resulting mixture wasstirred at room temperature for 3 h. The reaction solution was dilutedwith ether, washed successively with water and saturated sodium chloridesolution, dried over anhydrous magnesium sulfate, and concentrated invacuo. The residue was distilled under reduced pressure (110° C./3 torr)to obtain the benzyl derivative (6.65 g, 80%). Then, to a solution ofthis product (6.65 g, 29.9 mmol) dissolved in dichloromethane (30 ml),methanol (9 ml) and water (3 ml) was added trifluoroacetic acid (3 ml),and the mixture was stirred at room temperature for 3 h. After thereaction solution was neutralized with sodium hydroxide and condensed invacuo, the residue was purified by distillation under reduced pressureto obtain the title compound (4.71 g, 86%).

[0218]¹H-NMR (CDCl₃) δ: 7.32 (m, 5H, C₆H₅), 4.56 (s, 2H, CH₂Ph).

[0219] B L-1-O-Benzyl-2,3-di-O-docosyl-Sn-glycerol:

[0220] L-1-O-Benzyl-Sn-glycerol (1.90 g) and 1-bromo docosane (9.70 g)were reacted by the general procedure according to Example 15-A toobtain the title compound (7.0 g, 85%).

[0221]¹H-NMR (CDCl₃) δ: 7.30 (m, 5H, C₆H₅), 4.55 (s, 2H, CH₂Ph), 1.25(m, 76H, 38CH₂), 0.88 (t, 6H, J=6.6 Hz, 2CH₂CH₃).

[0222] C L-2,3-Di-O-docosyl-Sn-glycerol:

[0223] L-1-O-Benzyl-2,3-di-O-docosyl-Sn-glycerol (1.40 g) was reacted bythe general procedure according to Example 15-B to obtain the titlecompound (0.99 g, 79%) as white solid.

[0224]¹H-NMR (CDCl₃) δ: 1.25 (m, 76H, 38CH₂), 0.88 (t, 6H, J=6.6 Hz,2CH₂CH₃).

[0225] D3-O-[Methyl(5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)-onate]-L-2,3-di-O-docosyl-Sn-glycerol:

[0226] L-2,3-Di-O-docosyl-Sn-glycerol (990 mg) and methyl5-acetamido-4,7,8,9-tetra-O-acetyl-2-chloro-2,3,5-trideoxy-D-glycero-β-D-galacto-2-nonulopyranosonate[Chem. Ber., 99, 611 (1966)] (850 mg) were reacted by the generalprocedure according to Example 15-C to obtain the title compound (662mg, 40%) as white solid.

[0227]¹H-NMR (CDCl₃) δ: 3.79 (s, 3H, OCH₃), 2.60 (dd, 1H, J=4.8, 12.8Hz, H-3eq.), 2.15, 2.09, 2.04, 2.00, 2.00 (5s, 15H, 5Ac), 1.96 (t, 1H,J=12.8 Hz, H-3ax.), 1.25 (m, 76H, 38CH₂), 0.88 (t, 6H, J=6.6 Hz,2CH₂CH₃).

Example 431-O-[(5-Acetamido-3,5-dideoxy-D-glycero-α-D-galacto-nonulopyranosonicacid)-2-yl]-L-2,3-di-O-docosyl-Sn-glycerol

[0228]1-O-[Methyl(5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-L-2,3-di-O-docosyl-Sn-glycerol(632 mg) was reacted by the general procedure according to Example 16 toobtain the title compound (500 mg, 92%) as white solid.

[0229]¹H-NMR (CDCl₃-CD₃OD, 1:1, 40° C.) δ: 2.77 (dd, 1H, J=4.6, 13.4 Hz,H-3eq.), 2.02 (s, 3H, NAc), 1.74 (t, 1H, J=11.9 Hz, H-3ax.), 1.28 (m,76H, 38CH₂), 0.89 (t, 6H, J=6.8 Hz, 2CH₂CH₃).

Example 44 1-O-[Sodium{5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosyl}onate]-L-2,3-di-O-docosyl-Sn-glycerol

[0230]1-O-[(5-Acetamido-3,5-dideoxy-D-glycero-α-D-galacto-nonulopyranosonicacid)-2-yl]-L-2,3-di-O-docosyl-Sn-glycerol (200 mg) was reacted by thegeneral procedure according to Example 1 to obtain the title compound(237 mg, 83%) as white solid.

[0231]¹H-NMR (CD₃OD-D₂O, 1:1, 40° C.) δ: 2.93 (br.dd, 1H, H-3eq), 1.96(s, 3H, NAc), 1.77 (br.t, 1H, H-3ax), 1.28 (m, 76H, 38CH₂), 0.87 (t, 6H,J=6.6 Hz, 2CH₂CH₃).

Example 45 Methyl[2,2-bis(oleyl oxymethyl)propyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid]onate

[0232] A 2,2-Bis(Oleyl oxymethyl)propanol:

[0233] 1,1,1-Tris(hydroxymethyl)ethane (1.0 g) and oleyl chloride (5.25g) were reacted by the general procedure according to Example 15-A toobtain the title compound (2.91 g, 57%) as white solid.

[0234]¹H-NMR (CDCl₃) δ: 5.36 (m, 4H, 2CH═CH), 3.56 (d, 2H, J=5.9 Hz,CH₂OH), 1.27 (m, 40H, 20CH₂), 0.88 (t, 6H, J=6.6 Hz, 2CH₂CH₃), 0.85 (s,3H, CCH₃).

[0235] B Methyl[2,2-bis(oleyl oxymethyl)propyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid]onate:

[0236] 2,2-Bis(oleyl oxymethyl)propanol (400 mg) and methyl5-acetamido-4,7,8,9-tetra-O-acetyl-2-hloro-2,3,5-trideoxy-D-glycero-β-D-galacto-2-nonulopyranosonate[Chem. Ber., 99, 611 (1966)] (328 mg) were reacted by the generalprocedure according to Example 15-C to obtain the title compound (564mg, 80%) as white solid.

[0237]¹H-NMR (CDCl₃) δ: 5.35 (m, 4H, 2CH═CH), 3.78 (s, 3H, COOCH₃), 2.58(dd, 1H, J=5.0, 12.8 Hz, H-3eq.), 2.13, 2.12, 2.04, 2.02, 1.88 (5s, 15H,5Ac), 1.27 (m, 40H, 20CH₂), 0.90 (s, 3H, CCH₃), 0.88 (t, 6H, J=6.9 Hz,2CH₂CH₃).

Example 46 2,2-Bis(oleyl oxymethyl)propyl5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosidonic acid

[0238] Methyl[2,2-bis(oleyl oxymethyl)propyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid]onate(250 mg) was reacted by the general procedure according to Example 16 toobtain the title compound (190 mg, 89%) as white solid.

[0239]¹H-NMR (CDCl₃-CD₃OD, 1:1) δ: 5.35 (m, 4H, 2CH═CH), 2.76 (dd, 1H,J=4.2, 13.4 Hz, H-3eq), 2.05 (s, 3H, NAc), 1.70 (t, 1H, J=11.9 Hz,H-3ax), 1.30 (m, 40H, 20CH₂), 0.92 (s, 3H, CCH₃), 0.89 (t, 6H, J=6.8 Hz,2CH₂CH₃).

Example 47 Sodium[2,2-bis(oleyl oxymethyl)propyl5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosid)onate

[0240] 2,2-Bis(oleyl oxymethyl)propyl5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosidonic acid(913 mg) was reacted by the general procedure according to Example 1 toobtain the title compound (63 mg, 47%) as white solid.

[0241]¹H-NMR (CD₃OD-D₂O, 1:1, 50° C.) δ: 5.37 (m, 4H, 2CH═CH), 2.95 (dd,1H, J=5.3, 12.6 Hz, H-3eq), 1.97 (s, 3H, NAc), 1.74 (t, 1H, J=12.8 Hz,H-3ax), 1.28 (m, 40H, 20CH₂), 0.94 (s, 3H, CCH₃), 0.89 (t, 6H, J=6.8 Hz,2CH₂CH₃).

Example 48 Methyl[2,2-bis(docosyloxymethyl)butyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid]onate

[0242] A 2,2-Bis(docosyloxymethyl)butanol:

[0243] 1,1,1-Tris(hydroxymethyl)propane (1.0 g) and 1-bromo docosane(7.3 g) were reacted by the general procedure according to Example 15-Ato obtain the title compound (3.12 g, 56%) as white solid.

[0244]¹H-NMR (CDCl₃) δ: 3.59 (d, 2H, J=5.9 Hz, CH₂OH), 1.25 (m, 76H,38CH₂), 0.88 (t, 6H, J=6.7 Hz, 2CH₂CH₃), 0.84 (t, 3H, J=7.5 Hz,CCH₂CH₃).

[0245] B Methyl[2,2-bis(docosyloxymethyl)butyl5acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid]onate:

[0246] 2,2-Bis(docosyloxymethyl)butanol (2.26 g) and methyl5-acetamido-4,7,8,9-tetra-O-acetyl-2-chloro-2,3,5-trideoxy-D-glycero-β-D-galacto-2-nonulopyranosonate[Chem. Ber., 99, 611 (1966)] (1.02 g) were reacted by the generalprocedure according to Example 15-C to obtain the title compound (539mg, 22%) as white solid.

[0247]¹H-NMR (CDCl₃) δ: 3.78 (s, 3H, COOCH₃), 2.58 (dd, 1H, J=4.8, 12.8Hz, H-3eq.), 2.13, 2.12, 2.04, 2.02, 1.88 (5s, 15H, 5Ac), 1.95 (t, 1H,J=12.5 Hz, H-3ax.), 1.25 (m, 76H, 38CH₂), 0.88 (t, 6H, J=7.0 Hz,2CH₂CH₃), 0.84 (t, 3H, J=7.5 Hz, CCH₂CH₃).

Example 49 2,2-Bis(docosyloxymethyl)butyl5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosidonic acid

[0248] Methyl[2,2-bis(docosyloxymethyl)butyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosid]onate(520 mg) was reacted by the general procedure according to Example 16 toobtain the title compound (393 mg, 90%) as white solid.

[0249]¹H-NMR (CDCl₃-CD₃OD, 1:1, 60° C.) δ: 2.72 (dd, 1H, J=5.1, 12.1 Hz,H-3eq), 2.02 (s, 3H, NAc), 1.80 (t, 1H, J=11.9 Hz, H-3ax), 1.28 (m, 76H,38CH₂), 0.89 (t, 6H, J=7.0 Hz, 2CH₂CH₃), 0.86 (t, 3H, J=7.7 Hz,CCH₂CH₃).

Example 50 Sodium[2,2-bis(docosyloxymethyl)butyl5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosid)onate

[0250] 2,2-Bis(docosyloxymethyl)butyl5-acetamido-3,5-dideoxy-D-glycero-β-D-galacto-2-nonulopyranosidonic acid(388 mg) was reacted by the general procedure according to Example 1 toobtain the title compound (332 mg, 61%) as white solid.

[0251]¹H-NMR (CD₃OD-D₂O, 1:1, 60° C.) δ: 2.95 (br.dd, 1H, H-3eq), 1.96(s, 3H, NAc), 1.78 (br.t, 1H, H-3ax), 1.27 (m, 76H, 38CH₂), 0.87 (t, 9H,J=7.0 Hz, 3CH₂CH₃).

Example 513-O-[Methyl{5-N-(O-acetylglycolyl)-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl}onate]-1,2-di-O-docosyl-Sn-glycerol

[0252] [1,2-Di-O-docosyl-Sn-glycerol (400 mg) and methyl5-N-(O-acetylglycolyl)-4,7,8,9-tetra-O-acetyl-2-chloro-2,3,5-trideoxy-D-glycero-β-D-galacto-2-nonulopyranosonate[Carbohydr. Res., 174, 73 (1988)] (300 mg) were reacted by the generalprocedure according to Example 15-C to obtain the title compound (66 mg,10%) as white solid.

[0253]¹H-NMR (CDCl₃) δ: 4.60, 4.29 (2d, 2H, J=5.0 Hz, NHCOCH₂), 3.81 (s,3H, OCH₃), 2.62 (dd, 1H, J=4.8, 12.8 Hz, H-3eq), 2.20, 2.15, 2.13, 2.04,2.01 (5s, 15H, 5OAc), 1.25 (m, 76H, 38CH₂), 0.88 (t, 6H, J=7.0 Hz,2CH₂CH₃).

Example 523-O-[(5-N-glycolyl-3,5-dideoxy-D-glycero-α-D-galacto-nonulopyranosonicacid)-2-yl]-1,2-di-O-docosyl-Sn-glycerol

[0254]3-O-[Methyl{5-N-(O-acetylglycolyl)-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl}onate]-1,2-di-O-docosyl-Sn-glycerol(60 mg) was reacted by the general procedure according to Example 16 toobtain the title compound (43 mg, 88%) as white solid.

[0255]¹H-NMR (CDCl₃-CD₃OD, 1:1, 60° C.) δ: 2.78 (dd, 1H, J=4.0, 12.0 Hz,H-3eq), 1.79 (t, 1H, J=12.0 Hz, H-3ax), 1.28 (m, 76H, 38CH₂), 0.89 (t,6H, J=7.0 Hz, 2CH₂CH₃).

Example 53 3-O-[Sodium{5-N-(O-sodiumoxysulfonylglycolyl)-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosyl}onate]-1,2-di-O-docosyl-Sn-glycerol

[0256]3-O-[{-N-glycolyl-3,5-dideoxy-D-glycero-α-D-galacto-nonulopyranosonicacid}-2-yl]-1,2-di-O-docosyl-Sn-glycerol (41 mg) was reacted by thegeneral procedure according to Example 1 to obtain the title compound(16 mg, 26%) as white solid.

[0257]¹H-NMR (CD₃OD-D₂O, 1:1, 60° C.) δ: 2.99 (br.dd, 1H, H-3eq), 1.92(br.t, 1H, H-3ax), 1.30 (m, 76H, 38CH₂), 0.90 (t, 6H, J=7.0 Hz,2CH₂CH₃).

Example 543-S-[Methyl(5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-docosyl-Sn-thioglycerol

[0258] A 3-Bromo-3-deoxy-1,2-di-O-docosyl-Sn-glycerol:

[0259] 1,2-Di-O-docosyl-Sn-glycerol (200 mg, 0.28 mmol),N-bromosuccinimide (90 mg, 0.51 mmol) and triphenylphosphine (170 mg,0.65 mmol) were stirred in toluene (12 ml) at room temperature for 3days. The reaction solution was condensed in vacuo, and the residue waspurified by silica gel column chromatography (with 20 g of gel,hexane/toluene, 3:2) to obtain the title compound (168 mg, 77%) as whitepowder.

[0260]¹H-NMR (CDCl₃) δ: 1.25 (m, 76H, 38CH₂), 0.88 (t, 6H, J=7.0 Hz,2CH₂CH₃).

[0261] B3-S-[Methyl(5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-doosyl-Sn-thioglycerol:

[0262] Methyl5-acetamido-4,7,8,9-tetra-O-acetyl-2-S-acetyl-3,5-dideoxy-2-thio-D-glycero-α-D-galacto-2-nonulopyranosonate [J. Carbohydr. Chem., 5, 11(1986)] (114 mg, 0.21 mmol) and sodium methoxide (11 mg, 0.20 mmol) werestirred in anhydrous methanol (0.5 ml) at −10° C. for 1 h, and thereaction solution was evaporated to dryness in vacuo. To the residuewere added a solution of 3-bromo-3-deoxy-1,2-di-O-docosyl-Sn-glycerol(160 mg, 0.21 mmol) in dehydrated dimethylformamide (1.0 ml) and toluene(1.0 ml), and the mixture was stirred at room temperature for two days.The reaction solution was diluted with chloroform, washed with asaturated sodium chloride solution, and dried over anhydrous magnesiumsulfate. After the solvent was distilled off in vacuo, the residue werepurified by silica gel column chromatography (27 g of gel,toluene-acetone, 5:1) to obtain the title compound (85 mg, 34%) as whitepowder.

[0263]¹H-NMR (CDCl₃) δ: 3.79 (s, 3H, COOCH₃), 2.91 (dd, 1H, J=4.8, 13.0Hz, H-3eq.), 2.72 (m, 2H, SCH₂), 2.15, 2.13, 2.04, 2.03, 1.88 (5s, 15H,5Ac), 2.00 (t, 1H, J=12.8 Hz, H-3ax.), 1.25 (m, 76H, 38CH₂), 0.88 (t,6H, J=7.1 Hz, 2CH₂CH₃).

Example 553-S-[(5-Acetamido-3,5-dideoxy-D-glycero-α-D-galacto-nonulopyranosonicacid)-2-yl]-1,2-di-O-doosyl-Sn-thioglycerol

[0264]3-S-[Methyl(5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosyl)onate]-1,2-di-O-docosyl-Sn-thioglycerol(154 mg) was reacted by the general procedure according to Example 16 toobtain the title compound (115 mg, 86%) as white solid.

[0265]¹H-NMR (CDCl₃-CD₃OD, 1:1, 60° C.) δ: 2.94 (dd, 1H, J=4.6, 12.8 Hz,H-3eq), 2.84 (m, 2H, SCH₂), 2.02 (s, 3H, NAc), 1.84 (t, 1H, J=11.6 Hz,H-3ax), 1.29 (m, 76H, 38CH₂), 0.89 (t, 6H, J=6.9 Hz, 2CH₂CH₃).

Example 56 3-S-[Sodium{5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosyl}onate]-1,2-di-O-docosyl-Sn-thioglycerol

[0266]3-S-[(5-Acetamido-3,5-dideoxy-D-glycero-α-D-galacto-nonulopyranosonicacid)-2-yl]-1,2-di-O-doosyl-Sn-thioglycerol (422 mg) was reacted by thegeneral procedure according to Example 1 to obtain the title compound(29 mg, 45%) as white solid.

[0267]¹H-NMR (CD₃OD-D₂O, 1:1, 50° C.) δ: 3.06 (br.dd, 1H, H-3eq), 2.90(m, 2H, SCH₂), 1.90 (s, 3H, NAc), 1.30 (m, 76H, 38CH₂), 0.88 (t, 6H,J=7.0 Hz, 2CH₂CH₃).

Example 571-[N-(5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-2-O-methyl-D-glycero-β-D-galacto-2-nonulopyranosonyl)amino]-2,2-bis(docosyloxymethyl)propane

[0268] A 2,2-Bis(docosyloxymethyl)propyl methanesulfonate:

[0269] After a mixture of 2,2-bis(docosyloxymethyl)propanol (7.37 g; 10mmol), methanesulfonyl chloride (1.3 ml; 16.8 mmol) and pyridine (150ml) was stirred at 70° C. for 1 h, the reaction mixture was poured intowater. Precipitates thus obtained were collected by suction, washedsuccessively with water and acetone, and dried to obtain the titlecompound (6.66 g, 82%).

[0270]¹H-NMR (CDCl₃) δ: 4.13 (s, 2H, CH₂OMs), 2.98 (s, 3H, CH₃SO₂), 1.26(m, 76H, 38CH₂), 1.00 (s, 3H, CCH₃), 0.88 (t, 6H, J=7.0 Hz, 2CH₂CH₃).

[0271] B 1-Azido-2,2-bis(docosyloxymethyl)propane:

[0272] After a mixture of 2,2-bis(docosyloxymethyl)propylmethanesulonate (22.7 g; 27.8 mmol), sodium azide (0.27 g; 83.5 mmol),and dimethylformamide (150 ml) was stirred at 110° C. for 20 h, thereaction mixture was poured into water. Precipitates thus obtained werecollected by suction, washed successively with water and acetone, anddried to obtain the title compound (20.8 g; 98%).

[0273]¹H-NMR (CDCl₃) δ: 1.25 (m, 76H, 38CH₂), 0.89 (s, 3H, CCH₃), 0.88(t, 6H, J=7.0 Hz, 2CH₂CH₃).

[0274] C 1-Amino-2,2-bis(docosyloxymethyl)propane:

[0275] 1-Azido-2,2-bis(docosyloxymethyl)propane (12.8 g; 16.8 mmol) andpalladium hydroxide on carbon (6.0 g) in tetrahydrofuran (100 ml) wasstirred under the hydrogen atmosphere at 50° C. for 3 h. The reactionsolution was filtered through celite, and the filtrate was evaporated todryness in vacuo. The residue was purified by silica gel columnchromatography (60 g of gel, chloroform/methanol, 10:1) to obtain thetitle compound (8 g, 65%) as white powder.

[0276] D Methyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-β-D-galacto-2-nonulopyranosidonicacid:

[0277] After a mixture of methyl5-acetamido-3,5-dideoxy-D-glycero-β-D-galacto-2-nonulopyranosidonic acid[Chem. Ber., 99, 611 (1966)] (150 mg; 0.464 mmol), acetic anhydride (2.3ml) and pyridine (2.3 ml) was stirred at room temperature for 7 h, thereaction solution was evaporated to dryness in vacuo. The residue wasdissolved in chloroform, and treated with Amberist-15 (H type) to obtainthe title compound (218 mg, 96%) as white powder.

[0278]¹H-NMR (CDCl₃) δ: 3.33 (s, 3H, OCH₃), 2.56 (dd, 1H, J=4.8, 13.2Hz, H-3eq.), 2.16, 2.11, 2.06, 2.04, 1.91 (5s, 15H, 5Ac).

[0279] E1-[N-(5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-2-O-methyl-D-glycero-β-D-galacto-2-nonulopyranosonyl)amino]-2,2-bis(docosyloxymethyl)propane:

[0280] After a mixture of methyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-β-D-galacto-2-nonulopyranosidonicacid (260 mg; 0.53 mmol), 1-amino-2,2-bis(docosyloxymethyl)propane (390mg; 0.53 mmol), dicyclohexylcarbodiimide (165 mg; 0.80 mmol),1-hydroxybenztriazole (95 mg; 0.70 mmol) and chloroform (11 ml) wasstirred at room temperature for 17 h, the reaction solution was filteredthrough celite, an the solvent was distilled off in vacuo. The residuewas purified by silica gel column chromatography (with 60 g of gel,chloroform-acetone, 17:3) to obtain the title compound (620 mg, 97%) aswhite powder.

[0281]¹H-NMR (CDCl₃) δ: 3.18 (s, 3H, OCH₃), 2.51 (dd, 1H, J=4.8, 13.2Hz, H-3eq), 2.1 4, 2.08, 2.03, 2.01, 1.90 (5s, 15H, 5Ac), 1.77 (t, 1H,J=11.7 Hz, H-3ax), 1.25 (m, 76H, 38CH₂), 0.91 (s, 3H, CCH₃), 0.88 (t,6H, J=6.6 Hz, 2CH₂CH₃).

Example 581-[N-(5-Acetamido-3,5-dideoxy-2-O-methyl-D-glycero-β-D-galacto-2-nonulopyranosonyl)amino]-2,2-bis(docosyloxymethyl)propane

[0282] After a mixture of1-[N-(5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-2-O-methyl-D-glycero-β-D-galacto-2-nonulopyranosonyl)amino]-2,2-bis(docosyloxymethyl)propane(620 mg; 0.51 mmol), sodium methoxide (28 mg; 0.52 mmol), methanol (5ml) and tetrahydrofuran (5 ml) was stirred at room temperature for 7 h,the reaction solution was neutralized with Amberlist-15 (H type), andevaporated to dryness in vacuo. The residue was purified by silica gelcolumn chromatography (50 g of gel, chloroform/methanol, 10:1) to obtainthe title compound (480 mg, 90%) as white powder.

[0283]¹H-NMR (CDCl₃-CD₃OD, 1:1) δ: 3.24 (s, 3H, OCH₃), 2.40 (dd, 1H,J=4.9, 13.0 Hz, H-3eq), 2.05 (s, 3H, NAc), 1.27 (m, 76H, 38CH₂), 0.91(s, 3H, CCH₃), 0.89 (t, 6H, J=7.0 Hz, 2CH₂CH₃).

Example 591-[N-{5-Acetamido-3,5-dideoxy-2-O-methyl-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-β-D-galacto-2-nonulopyranosonyl}amino]-2,2-bis(docosyloxymethyl)propane

[0284]1-[N-(5-Acetamido-3,5-dideoxy-2-O-methyl-D-glycero-β-D-galacto-2-nonulopyranosonyl)amino]-2,2-bis(docosyloxymethyl)propane(831 mg) was reacted by the general procedure according to Example 1 toobtain the title compound (810 mg, 70%) as white solid.

[0285]¹H-NMR (CD₃OD-D₂O, 1:1, 40° C.) δ: 2.66 (br.dd, 1H, J=4.4, 12.5Hz, H-3eq), 2.03 (s, 3H, NAc), 1.79 (br.t, 1H, H-3ax), 1.31 (m, 76H,38CH₂), 0.91 (s, 3H, CCH₃), 0.91 (t, 6H, J=6.8 Hz, 2CH₂CH₃).

Example 601-[N-(5-Aetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-2-O-methyl-D-glycero-β-D-galacto-2-nonulopyranosonyl)amino]-2,2-bis(eicosyloxymethyl)propane

[0286] A 2,2-Bis(eicosyl oxymethyl)propyl methanesulfonate:

[0287] 2,2-Bis(eicosyloxymethyl)propanol (1.19 g) was reacted by thegeneral procedure according to Example 57-A to obtain the title compound(1.08 g, 82%) as white solid.

[0288]¹H-NMR (CDCl₃) δ: 4.13 (s, 2H, CH₂OMs), 2.98 (s, 3H, CH₃SO₂), 1.25(m, 68H, 34CH₂), 1.00 (s, 3H, CCH₃), 0.88 (t, 6H, J=7.0 Hz, 2CH₂CH₃).

[0289] B 1-Azido-2,2-bis(eicosyloxymethyl)propane:

[0290] 2,2-Bis(eicosyloxymethyl)propyl methanesulfonate (1.05 g) wasreacted by the general procedure according to Example 57-B to obtain thetitle compound (0.94 g, 96%) as white solid.

[0291]¹H-NMR (CDCl₃) δ: 1.25 (m, 68H, 34CH₂), 0.94 (s, 3H, CCH₃), 0.88(t, 6H, J=7.0 Hz, 2CH₂CH₃).

[0292] C 1-Amino-2,2-bis(eicosyloxymethyl)propane:

[0293] 1-Azido-2,2-bis(eicosyloxymethyl)propane (360 mg) was reacted bythe general procedure according to Example 57-C to obtain the titlecompound (206 mg, 59%) as white solid.

[0294]¹H-NMR (CDCl₃) δ: 2.74 (s, 2H, CH₂), 1.25 (m, 68H, 34CH₂), 0.90(s, 3H, CCH₃), 0.88 (t, 6H, J=7.0 Hz, 2CH₂CH₃).

[0295] D1-[N-(5-Acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-2-O-methyl-D-glycero-β-D-galacto-2-nonulopyranosonyl)amino]-2,2-bis(eicosyloxymethyl)propane:

[0296] Methyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-β-D-galacto-2-nonulopyranosidonicacid (179 mg) and 1-amino-2,2-bis(eicosyloxymethyl)propane (206 mg) werereacted by the general procedure according to Example 57-E to obtain thetitle compound (327 mg, 94%) as white solid.

[0297]¹H-NMR (CDCl₃) δ: 3.18 (s, 3H, OCH₃), 2.51 (dd, 1H, J=5.0, 13.0Hz, H-3eq), 2.14, 2.09, 2.04, 2.01, 1.91 (5s, 15H, 5Ac), 1.77 (dd, 1H,J=11.7, 12.8 Hz, H-3ax), 1.25 (m, 68H, 34CH₂), 0.91 (s, 3H, CCH₃), 0.88(t, 6H, J=7.0 Hz, 2CH₂CH₃).

Example 611-[N-(5-Acetamido-3,5-dideoxy-2-O-methyl-D-glycero-β-D-galacto-2-nonulopyranosonyl)amino]-2,2-bis(eicosyloxymethyl)propane

[0298]1-[N-(5-Acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-2-O-methyl-D-glycero-β-D-galacto-2-nonulopyranosonyl)amino]-2,2-bis(eicosyloxymethyl)propane(216 mg) was reacted by the general procedure according to Example 58 toobtain the title compound (202 mg, 74%) as white solid.

[0299]¹H-NMR (CDCl₃-CD₃OD, 10:1) δ: 3.22 (s, 3H, OCH₃), 2.36 (dd, 1H,J=4.8, 13.2 Hz, H-3eq), 2.04 (s, 3H, NAc), 1.26 (m, 68H, 34CH₂), 0.90(s, 3H, CCH₃), 0.88 (t,6H, J=7.1 Hz, 2CH₂CH₃).

Example 621-[N-{5-Aetamido-3,5-dideoxy-2-O-methyl-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-β-D-galacto-2-nonulopyranosonyl}amino]-2,2-bis(eicosyloxymethyl)propane

[0300]1-[N-(5-Acetamido-3,5-dideoxy-2-O-methyl-D-glycero-β-D-galacto-2-nonulopyranosonyl)amino]-2,2-bis(eicosyloxymethyl)propane(422 mg) was reacted by the general procedure according to Example 1 toobtain the title compound (49 mg, 36%) as white solid.

[0301]¹H-NMR (CD₃OD-D₂O, 1:1, 40° C.) δ: 2.66 (br.dd, 1H, H-3eq), 2.02(s, 3H, NAc), 1.78 (br.t, 1H, J=12.3 Hz, H-3ax), 1.30 (m, 68H, 34CH₂),0.90 (s, 3H, CCH₃), 0.90 (t, 6H, J=7.0 Hz, 2CH₂CH₃).

Example 631-[N-(5-Acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-2-O-ethyl-D-glycero-β-D-galacto-2-nonulopyranosonyl)amino]-2,2-bis(docosyloxymethyl)propane

[0302] A Methyl(ethyl5-acetamido-3,5-dideoxy-D-glycero-β-D-galacto-2-nonulopyranosid)onate:

[0303] A mixture of N-acetylneuraminic acid (1.00 g; 3.23 mmol),Dowex-50 (H form) (15 g) and dehydrated ethanol (100 ml) was heated atreflux for 18 h. The reaction mixture was packed into a column, andeluted with 2N-HCl-methanol (100 ml). After the solvent was distilledoff in vacuo. The residue thus obtained was stirred with aceticanhydride (20 ml) and pyridine (20 ml) at room temperature for 17 h, andthen the reaction solution was evaporated to dryness in vacuo. Theresidue was dissolved in chloroform, washed successively with 0.1N HCl,water and saturated NaCl solution, dried over anhydrous magnesiumsulfate, and then the solvent was distilled off in vacuo. After theresidue thus obtained was stirred with sodium methoxide (31 mg; 0.575mmol) in methanol (23 ml) at room temperature for 17 h, the reactionmixture was neutralized with Dowex-50 (H form), and then the solvent wasdistilled off. The residue was purified by silica gel columnchromatography (40 g of gel, chloroform/methanol, 4:1) to obtain thetitle compound (142 mg, 12%) as white powder.

[0304]¹H-NMR (CD₃OD) δ: 3.78 (s, 3H, OCH₃), 2.36 (dd, 1H, J=4.9, 13.0Hz, H-3eq.), 2.00 (s, 3H, NAc), 1.62 (dd, 1H, J=11.4, 12.8 Hz, H-3ax.),1.16 (t, 3H, J=7.0 Hz, CH₂CH₃).

[0305] B Methyl(ethyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-β-D-galacto-2-nonulopyranosid)onate:

[0306] Methyl(ethyl5-acetamido-3,5-dideoxy-D-glycero-β-D-galacto-2-nonulopyranosid)onate(142 mg) was reacted by the general procedure according to Example 57-Dto obtain the title compound (209 mg, 99%) as white solid.

[0307]¹H-NMR (CDCl₃) δ: 3.80 (s, 3H, COOCH₃), 2.45 (dd, 1H, J=5.1, 12.8Hz, H-3eq.), 2.15, 2.08, 2.03, 2.02, 1.89 (5s, 15H, 5Ac), 1.87 (dd, 1H,J=11.4, 12.8 Hz, H-3ax.), 1.2 (t, 3H, J=7.1 Hz, CH₂CH₃).

[0308] C Ethyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-β-D-galacto-2-nonulopyranosidonicacid:

[0309] A mixture of methyl (ethyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-β-D-galacto-2-nonulopyranosid)onate(203 mg, 0.39 mmol), anhydrous lithium iodide (523 mg, 3.9 mmol) andpyridine (8.0 ml) was stirred at 90° C. for 14 h. The reaction solutionwas subjected to gel filtration (LH-20, 150 ml, methanol), and thenpurified by silica gel column chromatography (20 g of gel,chloroform/methanol, 7:3) to obtain the title compound (92 mg, 47%) aswhite solid.

[0310]¹H-NMR (CDCl₃) δ: 3.52 (m, 2H, CH₂CH₃), 2.56 (dd, 1H, J=4.5, 12.2Hz, H-3eq.), 2.15, 2.11, 2.06, 2.04, 1.91 (5s, 15H, 5Ac), 1.92 (t, 1H,J=11.8 Hz, H-3ax.), 1.24 (t, 3H, J=7.0 Hz, CH₂CH₃).

[0311] D1-[N-(5-Acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-2-O-ethyl-D-glycero-β-D-galacto-2-nonulopyranosonyl)amino]-2,2-bis(docosyloxymethyl)propane:

[0312] Ethyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-β-D-galacto-2-nonulopyranosodonicacid (92 mg) and 1-amino-2,2-bis(docosyloxymethyl)propane (268 mg) werereacted by the general procedure according to Example 57-E to obtain thetitle compound (191 mg, 86%) as white solid.

[0313]¹H-NMR (CDCl₃) δ: 2.52 (dd, 1H, J=4.8, 13.2 Hz, H-3eq), 2.14,2.08, 2.03, 2.01, 1.91 (5s, 15H, 5Ac), 1.76 (t, 1H, J=12.4 Hz, H-3ax),1.25 (m, 76H, 38CH₂), 1.19 (t, 3H, J=7.0 Hz, OCH₂CH₃), 0.90 (s, 3H,CCH₃), 0.88 (t, 6H, J=7.0 Hz, 2CH₂CH₃).

Example 641-[N-(5-Acetamido-3,5-dideoxy-2-O-ethyl-D-glycero-β-D-galacto-2-nonulopyranosonyl)amino]-2,2-bis(docosyloxymethyl)propane

[0314]1-[N-(5-Acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-2-O-ethyl-D-glycero-β-D-galacto-2-nonulopyranosonyl)amino]-2,2-bis(docosyloxymethyl)propane(185 mg) was reacted by the general procedure according to Example 58 toobtain the title compound (114 mg, 71%) as white solid.

[0315]¹H-NMR (CDCl₃-CD₃OD, 10:1) δ: 2.38 (dd, 1H, J=4.8, 13.2 Hz,H-3eq), 2.04 (s, 3H, NAc), 1.26 (m, 76H, 38CH₂), 1.17 (t, 3H, J=7.0 Hz,OCH₂CH₃), 0.89 (s, 3H, CCH₃), 0.88 (t, 6H, J=7.3 Hz, 2CH₂CH₃).

Example 651-[N-{5-Acetamido-3,5-dideoxy-2-O-ethyl-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-β-D-galacto-2-nonulopyranosonyl}amino]-2,2-bis(docosyloxymethyl)propane

[0316]1-[N-(5-Acetamido-3,5-dideoxy-2-O-ethyl-D-glycero-β-D-galacto-2-nonulopyranosonyl)amino]-2,2-bis(docosyloxymethyl)propane(422 mg) was reacted by the general procedure according to Example 1 toobtain the title compound (140 mg, 93%) as white solid.

[0317]¹H-NMR (CD₃OD-D₂O, 1:1, 40° C.) δ: 2.67 (br.dd, 1H, H-3eq), 2.02(s, 3H, NAc), 1.77 (br.t, 1H, H-3ax), 1.30 (m, 76H, 38CH₂), 1.25 (t, 3H,J=7.0 Hz, OCH₂CH₃), 0.91 (t, 6H, J=6.8 Hz, 2CH₂CH₃), 0.90 (s, 3H, CCH₃).

Example 661-[N-(5-Acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-2-O-methyl-D-glycero-α-D-galacto-2-nonulopyranosonyl)amino]-2,2-bis(docosyloxymethyl)propane

[0318] A Methyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosidonicacid:

[0319] Methyl5-acetamido-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosidonic acid[Chem. Ber., 99, 611 (1966)] (150 mg) was reacted by the generalprocedure according to Example 57-D to obtain the title compound (220mg, 96%) as white solid.

[0320]¹H-NMR (CDCl₃-CD₃OD, 1:1) δ: 3.20 (s, 3H, OCH₃), 2.42 (dd, 1H,J=4.4, 12.5 Hz, H-3eq.), 1.96, 1.95, 1.87, 1.83, 1.70 (5s, 15H, 5Ac),1.56 (t, 1H, J=12.5 Hz, H3ax.).

[0321] B1-[N-(5-Acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-2-O-methyl-D-glycero-α-D-galacto-2-nonulopyranosonyl)amino]-2,2-bis(docosyloxymethyl)propane:

[0322] Methyl5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-α-D-galacto-2-nonulopyranosidonicacid (191 mg) and 1-amino-2,2-bis(docosyloxymethyl)propane (396 mg) werereacted by the general procedure according to Example 57-E to obtain thetitle compound (345 mg, 73%) as white solid.

[0323]¹H-NMR (CDCl₃) δ: 3.36 (s, 3H, OCH₃), 2.13, 2.08, 2.03, 2.01, 1.89(5s, 15H, 5Ac), 1.98 (t, 1H, J=11.7 Hz, H3ax.) 1.25 (m, 76H, 38CH₂),0.90 (s, 3H, CCH₃), 0.88 (t, 6H, J=7.0 Hz, 2CH₂CH₃).

Example 671-[N-(5-Acetamido-3,5-dideoxy-2-O-methyl-D-glycero-α-D-galacto-2-nonulopyranosonyl)amino]-2,2-bis(docosyloxymethyl)propane

[0324]1-[N-(5-Acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-2-O-methyl-D-glycero-α-D-galacto-2-nonulopyranosonyl)amino]-2,2-bis(doosyloxymethyl)propane(340 mg) was reacted by the general procedure according to Example 58 toobtain the title compound (216 mg, 74%) as white solid.

[0325]¹H-NMR (CDCl₃-CD₃OD, 10:1) δ: 3.36 (s, 3H, OCH₃), 2.55 (dd, 1H,J=4.8, 13.2 Hz, H-3eq), 2.03 (s, 3H, NAc), 1.82 (dd, 1H, J=11.2, 13.0Hz, H3ax.), 1.26 (m, 76H, 38CH₂), 0.91 (s, 3H, CCH₃), 0.88 (t, 6H, J=7.0Hz, 2CH₂CH₃).

Example 681-[N-{5-Acetamido-3,5-dideoxy-2-O-methyl-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosonyl}amino]-2,2-bis(docosyloxymethyl)propane

[0326]1-[N-(5-Acetamido-3,5-dideoxy-2-O-methyl-D-glycero-α-D-galacto-2-nonulopyranosonyl)amino]-2,2-bis(docosyloxymethyl)propane(117 mg) was reacted by the general procedure according to Example 1 toobtain the title compound (98 mg, 60%) as white solid.

[0327]¹H-MR (CD₃OD-D₂O, 1:1, 40° C.) δ: 2.55 (br.dd, 1H, J=5.1, 13.6 Hz,H-3eq), 1.99 (s, 3H, NAc), 1.30 (m, 76H, 38CH₂), 0.90 (br.t, 9H, J=6.6Hz, CCH₃ and 2CH₂CH₃).

[0328] Assessment of Growth Inhibitory Effect on HIV-1 and Cytotoxicity

[0329] Samples of various concentrations (25 μl) are distributed into a24-well microplate, immediately added with human T cells (H9 cell line,1.2×10⁵/ml) (1 ml), covered, and incubated at 37° C. for 1 h. Then, AIDSvirus (HIV-1, IIIb, 100 TCID 50) (25 μl) is added to each well andthoroughly mixed, covered again, and incubated under the CO₂ atmosphereat 37° C. for 3 days. On the fourth day, aliquots (200 μl) of theculture medium from each well are transferred into a fresh microplate.The growth medium (500 μl) is added to each well, and further incubatedfor 2 days.

[0330] On the 6th day, aliquots (90 μl) of the culture supernatant arewithdrawn, and assayed the amount of p24, a constituent of HIV-1, byELISA method using the anti-p24 antibody to assess the growth of saidvirus. Also, using the remaining cell culture suspension (100 μl),viable cells are counted by the MTT method. Experimental values areexpressed as the ratios (%) to the negative control value (measuredusing the solvent to dissolve test compounds as the sample) to representthe anti-HIV activity and cytoxicity. Then, for the unifiedrepresentation of the anti-HIV activity and cytotoxicity, theconcentrations of test compounds at their 50% values are obtained fromthe activity-concentration curves and the cytotoxicity-concentration,and represented as IC50 and CC50, respectively.

[0331] Results of the Anti-HIV Activity Assay

[0332] In the following table are shown the compounds and theirstructural characteristics having IC50 (μM) values less than 100 μM.Carbon number of the functional group (in parentheses) represents thatof alkyl group of lipid side-chain beyond the ether linkage, and,particularly, in the case where lipid is alkylglycerol, that of alkylgroup of said alkyl glycerol. TABLE 1 Example No. IC 50 (μM) Functionalgroup (carbon number) 5 46.0 Two alkyl chains (10) 6 50.0 Two alkylchains (10), β-isomer of 5 7 16.0 Two alkyl chains (14) 8 15.0 Two alkylchains (14) β-isomer of 7 9 3.8 Two alkyl chains (18) 10 30.0 One alkylchain (18), with a sulfate group as a side chain 11 13.0 One alkyl chain(18), β-isomer of 10 12 0.8 Two alkyl chains (22) 13 1.6 Two alkylchains (22), β-isomer of 12 14 15.0 Two alkyl chains (14), thioglycoside17 2.5 Two alkyl chains (20) 20 1.2 Two alkyl chains (24) 24 34.0Cholesterol 25 100.0 One alkyl chain (18), unsaturated 26 100.0 Onealkyl chain (18) 29 1.6 Two alkyl chains (22), branched 32 2.5 Two alkylchains (20), branched 35 90.0 Aromatic 38 2.5 Two alkyl chains (22),glycero-ester 41 4.0 Two alkyl chains (22), KDN 50 1.1 Two alkyl chains(22), with an ethyl group at the branching point 59 1.4 Two alkyl chains(22), amide linkage

[0333] These results indicate that, as the lipid bound to nonulonic acid(sialic acid and KDN) via glycosidic linkage are preferred alkyl,carbonyloxy or oxycarbonyl groups. Furthermore, no significantdifference observed in the activity of Example 7 (O-glycoside) ascompared with that of Example 14 (S-glycoside) indicates that thelinkage between carbohydrate and lipid moieties can be eitherO-glycosidic or S-glycosidic. Also, no difference observed in theactivity between Example 29 and Example 59, wherein they differ only inthat the former has the O-glycosidic linkage and the latter the amidelinkage, leads to the conclusion that the linkage between carbohydrateand lipid can be of any form.

[0334] So far as sulfuric acid derivatives of sialoglycerolipids areconcerned, as evidently seen comparing Example 9 (two alkyl chains, eachhaving 18 carbon atoms, IC50: 3.8) and Example 11 (one alkyl chain with18 carbon atoms, IC50: 13.0), both derivatives, one with single chain,and the other with two chains, have the enough activity for thepractical use, but the latter one is generally more active. Therefore,derivatives with two alkyl chains can be more preferable than those withone alkyl chain.

[0335] Also, as evidently seen comparing Example 38 (two alkyl chains,each having 22 carbon atoms, glyceroester, IC50: 2.5) and Example 12(two alkyl chains, each having 22 carbon atoms, IC50: 0.8), thesecompounds have high activity regardless of whether their glycero-lipidmoieties are either alkylglycerol or acylglycerol. Therefore, thebiological activity is not so much affected by the linkage form betweenthe glycerol skeleton and the long alkyl chain group, but rather mainlydependent on the length of long alkyl chain group.

[0336] So far as compounds wherein glycerol having two alkyl chains islinked to sugar via glycosidic linkage are concerned, as shown in Table1, minimum IC50 values are associated mostly with those with carbonatoms of 18 (Example 9), 20 (Example 17), 22 (Example 12) and 24(Example 20). Therefore, in the case where the sulfate derivatives ofsialoglycerolipids related to this invention are either alkyl glycerolor acyl glycerol, number of carbon atoms of alkyl groups linked to theglycerol moiety is thought to be preferably 18˜24. Since the biologicalactivity depends mainly on the length of long-chain alkyl group linkedto the glycerol skeleton, said long chain alkyl group can be saturatedor unsaturated, also branched. In the case of alkyl glycerol with 24carbon atoms as in Example 20, the “number of skeleton-forming atoms”per one side chain amounts to 25. This is because the “number ofskeleton-forming atoms” includes the oxygen atom related to the etherlinkage, in addition to carbon atoms of alkyl group. Furthermore, in thecase of pseudo-glycerol as in Example 29 and Example 32, since thecarbon atom of methylene group between the carbon atom at the β positionof the main chain of glycerol moiety and the oxygen atom of the etherlinkage is also included in the “number of skeleton-forming atoms”, thatnumber becomes 24 in the case of Example 29, and 22 in the case ofExample 32.

[0337] Also, the glycerol moiety can have a methyl branch, and amethylene group between the main chain of the glycerol moiety and etherlinkage or ester linkage (Example 29 and Example 32). Therefore, theglycerol moiety can have a short alkyl chain such as methyl, ethyl,propyl, butyl groups, etc. at the β carbon atom of its main chain (inthis case, however, these groups cannot be called side chain or branchedchain because of insufficient length of alkyl groups.), and one to threecarbon atoms can be present between the main chain of glycerol moietyand the ether or ester linkage.

[0338] In comparison of Example 29 with Example 12, two chains extendingfrom the β carbon atom of the main chain of glycerol moiety are of thesame length in Example 29, while they are of the different length inExample 12. That is, in Example 29, number of “skeleton-forming atom”per one chain of the two chains extending from the β carbon atom of themain chain of glycerol moiety is the same 24, but different in Example12, as 23 and 24. Therefore, it can be concluded that, in the case oftwo chains, number of “skeleton-forming atoms” per one chain,furthermore, number of alkyl chain-forming carbon atoms can bedifferent. In addition, as to Example 29, it is advantageous that thetwo chains are completely the same having the same component atoms andconfiguration because for the easy preparation.

[0339] In the table below are shown CC50s. TABLE 2 Example No. CC 50(μM) Example No. CC 50 (μM) 5 >200 20 >20 6 >200 24 >200 7 >200 25 >2008 >200 26 >200 9 >200 29 >20 10 >200 32 >200 11 200 35 >20 12 >20038 >200 13 >200 41 >200 14 >200 50 >200 17 >20 59 >200

[0340] These results indicate that compounds related to the presentinvention are generally low in the cytotoxicity.

[0341] Assay of Antiviral Activity against other Viruses than HIV

[0342] Antiviral activity against human Parainfluenza virus, Respiratorysyncytial virus (RSV) and Herpes simplex II virus (HSVII) was determinedby the plaque reduction assay, while that against Felineimmunodeficiency virus (FIV) and Feline leukemia virus (FeLV) by thereverse transcriptase assay and ELISA, respectively.

[0343] 1) Plaque Reduction Assay

[0344] a) Preparation viruse, host cell and virus stock

[0345] Each virus was obtained from ATCC. As host cells, VERO cells wereused for Parainfluenza virus, and HEP2 cells for other viruses.

[0346] To fibroblasts growing in a T150 flask were added virus-infectedcells, and the cells were incubated until they reached 60-80% infection.Cells were trypsinized, recovered and used as the virus stock. As thecell culture medium was used E-MEM containing 2-5% FBS, 100 U/mlPenicillin, 2.5 μg/ml Amphotericin and 10 μg/ml Gentamycin.

[0347] b) Preparation of sample

[0348] Compound related to Example 29 was appropriately diluted to theconcentrations of 500, 100, 50, 10 and 5 μg/ml with the culture mediumor methyl cellulose. As the reference drug for HSV II was usedAcyclovir.

[0349] c) Assay procedure

[0350] Anti-viral Activity

[0351] After cells were cultured in monolayer in 24-well microplates,the supernatant was discarded, replaced with a diluted sample (compound)solution (0.5 ml), and the mixture was incubated at 36-38° C. under the5-7% CO₂ atmosphere for 1 h. At the same time, to the cell control welland virus control wells was added the equal amount of medium.

[0352] After the supernatant was discarded, the medium (0.5 ml) and thevirus stock (0.2 ml) were added to the sample well and virus well,respectively. After incubating again for 1 h, the sample solution (1 ml)diluted with methylcellulose containing FBS was overlayered. To thecontrol well was added only the same amount of the vehicle.

[0353] Cells were cultured at 36-38° C. under the 5-7% CO₂ atmosphere,and examined for the appearance of plaques under microscope. Monolayerof cells was fixed with 10% formalin, washed with water, stained with0.8% crystal violet, and dried. Number of plaques counted in the samplewell was compared with that in the virus control well, and the decreasein the number of plaques was used to express the antiviral activity.

[0354] Cytotoxicity Assay

[0355] Of the anti-viral activity assay procedures mentioned above, thevirus infection step was omitted from sample wells, and cells werecultured in the presence of samples at various concentrations, andviable cell number was compared with that in the cell control well.Surviving cell number was counted by the tetrazolium method for themicroplate assay.

[0356] 2) Reverse Transcriptase Assay

[0357] a) Preparation of host cell and virus

[0358] Each virus was obtained from ATCC. FIV was cultured with CRFKcells in Eagle's balanced salt medium containing 10% FBS, and thesupernatant was used as the virus stock. As FeLV was used ATCC VR-717.

[0359] b) Preparation of sample

[0360] Compound related to Example 29 was diluted with the medium tosimilar concentrations described above.

[0361] c) Assay procedure

[0362] For each single strain of virus, the assay was performed in thefollowing combinations of conditions. Medium control well Culture mediumCell control well Culture medium + cell Cytotoxicity well Culturemedium + cell + sample Virus control well Culture medium + cell + virustest well Culture medium + cell + virus + sample Color control wellCulture medium + sample

[0363] After CRFL cells were cultured in monolayer in 96-wellmicroplates, the supernatant was discarded. Then, to the medium controlwell, the cell control well and the virus control well was added themedium (0.2 ml), and to the cytotoxicity well, the test well and thecolor control well was added the diluted sample solution at 0.2, 0.1,and 0.2 ml, respectively, and incubated under the 5-7% CO₂ atmosphere at36-38° C. for 1 h. Then, supernatants of the virus control well and thetest well were discarded, and the virus stock (0.1 ml) was added, andincubated further for 1 h. The, after supernatants were removed, and, tothe virus control well was added the medium (0.2 ml), and to the testwell the same volume of the diluted sample, then the incubation wasfurther continued.

[0364] Reverse Transcriptase Assay by ELISA

[0365] Using supernatants as the sample from the cell control well, thevirus control well and the test well, the reverse transcriptase of FIVor FeLV antigen contained therein were quantitated using assay kits fromAmersham LIFESCIENCE and Synbiotics, respectively. Anti-viral activitywas expressed as that ratio (%) of the value of the test well to that ofthe virus control well after both values were subtracted with the valueof the cell control well, respectively.

[0366] Cytotoxicity Assay

[0367] Viable cell numbers in the cytotoxicity well and the cell controlwell were counted by the tetrazolium method for microplate assaysimilarly as described above, and the cytotoxicity was expressed as theratio (%) of the cell number of the former to that of the latter.

[0368] Anti-viral Activity other than HIV and Cytotoxicity

[0369] Results of anti-viral activity assay are shown in the tablebelow. TABLE 3 Anti-viral activity of compound related to Example 29IC50 (μM) Virus Example 29 Acyclovir Herpes simplex II virus 45.2 21.6Respiratory syncytial virus 5.1 — Parainfluenza virus 37.6 — Felineimmunodeficiency virus 4.5 — Feline leukemia virus 58.3 —

[0370] These results indicate that the compound related to thisinvention has not only the anti-HIV activity but also the anti-viralactivity against other viruses.

[0371] Furthermore, when CC50s were calculated for each virus by thesimilar method as described above, they all exceeded the value 500 μM.Compounds related to this invention have the anti-viral activity againstother viruses and low cytotoxicity, indicating that they are not onlyeffective in treating various diseases but also highly safe in theliving body and effective as therapeutics.

[0372] Anti-coagulation Activity

[0373] To the human plasma (400 μl) were added compound diluted withphysiological saline (500 μl) followed by 2% calcium chloride solution(100 μl), and the mixture was incubated at 37° C. for 30 min todetermine the minimum concentration (μM) of said compound to exhibit theanti-coagulant activity. Plasma coagulation was judged with the nakedeye.

[0374] Toxicity Study

[0375] Compounds were intravenously administered to mice, and LD50s(mg/kg) were determined.

[0376] Results of Anticoagulant Activity Assay and Toxicity Study TABLE4 Minimum concentration exhibiting anticoagulant action (μM) LD50(mg/kg) Dextran sulfate 0.5 — Example 12 900 2000 Example 13 1200 2000Example 29 900 2000

[0377] These results indicate that compounds related to this inventionare generally low not only in the cytotoxicity but also in theanticoagulant action, especially with compounds related to Examples 12,13 and 29, furthermore they are low in the toxicity compounds related toExamples 12, 13 and 29, furthermore they are low in the toxicity to theliving body. Therefore, it is obvious that pharmaceutics containingcompounds related to this invention at the effective doses arepreferable as drugs.

[0378] Effective dose of compounds related to this invention or thesalts thereof can be determined by the general method well known tothose skilled in the art including the method of establishing thedose-reaction curve in appropriate animal model or non-human primate andextrapolating its data to humans, or that of determining the dose in theclinical test.

[0379] Preferable doses of drugs, anti-viral agents and anti-HIV agentsrelated to this invention are varied by various factors such as theseverity of disease, body weight and age of individuals, half life ofdrug in the blood circulation, etc., and they can be easily determinedby those skilled in the art.

[0380] Medicine related to this invention can be administered by variousways such as intravenous injection, oral administration, inhalation,etc. Pharmaceutical carriers, diluents and excipients can be easilyselected by those skilled in the art according to the clinical use ofdrug, and, if necessary, supplements such as disintegrator, binder(including liposome), surfactant, emulsifier, buffer, solubilizing agentor preservative are added to make liquid preparation, emulsion orsuspension.

[0381] As described above, novel compounds related to this inventionhave significant effectiveness such as not only the high antiviralactivity but also the low cytotoxicity. Therefore, novel compoundsrelated to the present invention are optimal as the antiviral agent.

[0382] Also, since novel compounds related to the present invention arelow not only in the anticoagulant action but also in the toxicity to theliving body, pharmaceutical preparations containing them in effectivedoses are preferable as medicine. In view of the bleeding tendencyobserved particularly with HIV patients, it is obvious that these novelcompounds are extremely useful as the anti-HIV medicine.

What is claimed is:
 1. A method of treating a virus-mediated disease ina mammal comprising administering to said mammal a sulfated sialyl lipidcompound, or the salt thereof wherein said sulfated sialyl lipidcompound or salt thereof comprises: a monosaccharide moiety linked to alipid moiety, wherein said monosaccharide moiety is a sialic acid or2-keto-3-deoxy-D-glycero-2-nonuloic acid (KDN), and said lipid moiety islinked to the anomeric carbon of said sialic acid or said KDN, and allhydroxyl groups of said sialic acid or said KDN are sulfated.
 2. Themethod of treating a virus-mediated disease in a mammal according toclaim 1, wherein The link between said sialic acid or said KDN of saidmonosaccharide moiety and said lipid moiety is an O-glycosidic linkageor an S-glycosidic linkage at position 2 of said sialic acid or saidKDN, or an amide linkage at position 1 of said sialic acid or said KDN.3. The method of treating a virus-mediated disease in a mammal accordingto claim 2, wherein said lipid moiety is a linear lipid comprising abranched chain structure.
 4. The method of treating a virus-mediateddisease in a mammal according to claim 3, wherein said lipid moietycomprises a branched chain structure, wherein a site of said branchedchain structure is located at position 2 of a main chain of said lipidmoiety.
 5. The method of treating a virus-mediated disease in a mammalaccording to claim 4, wherein said branched chain structure of saidlipid moiety comprises two chains due to said branching.
 6. The methodof treating a virus-mediated disease in a mammal according to claim 5,wherein said lipid moiety bas an alkyl group with 1 to 4skeleton-forming atoms at the site of said branching.
 7. The method oftreating a virus-mediated disease in a mammal according to claim 6,wherein the total number of skeleton-forming atoms of said lipid moietyis 22 to
 60. 8. The method of treating a virus-mediated disease in amammal according to claim 7, wherein said branched chains comprise anunsaturated bond between carbon atoms.
 9. The method of treating avirus-mediated disease in a mammal according to claim 7, wherein saidbranched chains of said lipid moiety are linear.
 10. The method oftreating a virus-mediated disease in a mammal according to claim 9,wherein said branched chains comprise an ester linkage or an etherlinkage.
 11. The method of treating a virus-mediated disease in a mammalaccording to claim 10, wherein said ester linkage or said ether linkageis localized at position 1 or 2 of said branched chain structure. 12.The method of treating a virus-mediated disease in a mammal according toclaim 7, wherein the number of skeleton-forming atoms per one branchedchain is 10 to
 28. 13. The method of treating a virus-mediated diseasein a mammal according to claim 12, wherein the number ofskeleton-forming atoms per one branched chain is 18 to
 26. 14. Themethod of treating a virus-mediated disease in a mammal according toclaim 13, wherein the number of skeleton-forming atoms per one branchedchain is
 24. 15. The method of treating a virus-mediated disease in amammal according to claim 14, wherein said branched chains are of thesame length, respectively.
 16. The method of treating a virus-mediateddisease in a mammal according to claim 15, wherein said branched chainsare of the same structure, respectively.
 17. The method of treating avirus-mediated disease in a mammal according to claim 16, wherein saidbranched chains have an ester linkage or an ether linkage at position 1or 2 of said branched chains.
 18. The method of treating avirus-mediated disease in a mammal according to claim 17, wherein saidsulfated sialyl lipid compound is sodium(2,2-bis(docosyloxymethyl)propyl5-acetamido-3,5-dideoxy4,7,8,9-tetra-O-sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosid)onate or the acidthereof.
 19. The method of treating a virus-mediated disease in a mammalaccording to claim 18, wherein one or more sodium atoms is replaced by adifferent cation.
 20. The method of treating a virus-mediated disease ina mammal according to claim 19, wherein said different cation ispotassium.
 21. The method according to any one of claims 1-20, whereinsaid sulfated sialyl lipid compound is administered to said mammal by aroute selected from the group consisting of an intravenous route, anintramuscular route, an intraperitoneal route, an intra-arterial route,a subcutaneous route, a percutaneous route, a sublingual route, aninhalation route, and a rectal route.
 22. The method of treating avirus-mediated disease in a mammal according to any one of claims 1-20,wherein said virus is selected from the group consisting of a humanimmunodeficiency virus, a feline immunodeficiency virus, a felineleukemia virus, a herpes simplex II virus, a parainfluenza virus, arespiratory syncytial virus, and combinations thereof.
 23. The method oftreating a virus-mediated disease in a mammal according to claim 22,wherein said virus is a respiratory syncytial virus.
 24. A compound orthe salt thereof which is a glycoside comprising a monosaccharide-lipid,wherein said lipid moiety is linked to the anomeric carbon of sialicacid or KDN, and all the hydroxyl groups of said sialic acid moiety orKDN moiety are sulfated, wherein said compound issodium(2,2-bis(docosyloxymethyl)propyl5-acetamido-3,5-dideoxy-4,7,8,9-tetra-O-(sodiumoxysulfonyl)-D-glycero-α-D-galacto-2-nonulopyranosid)onate or an acidform thereof.
 25. The compound according to claim 24, wherein one ormore sodium atoms is replaced by a different cation.
 26. The compoundaccording to claim 25, wherein said different cation is potassium.
 27. Apharmaceutical composition comprising a compound according to claim 24and a pharmaceutically acceptable carrier.
 28. A pharmaceuticalcomposition comprising a compound according to claim 25 and apharmaceutically acceptable carrier.
 29. A pharmaceutical compositioncomprising a compound according to claim 26 and a pharmaceuticallyacceptable carrier.